Gene linked to osteoarthritis

ABSTRACT

A role of the human MATN3 gene in osteoarthritis is disclosed. Methods for diagnosis, prediction of clinical course and treatment for osteoarthritis using polymorphisms in the MATN3 gene are also disclosed.

RELATED APPLICATIONS

[0001] This application claims the benefit of U.S. application Ser. No.60/431,538 filed on Dec. 5, 2002 and also claims the benefit of U.S.application Ser. No. 10/057,312 filed on Jan. 25, 2002 (converted toProvisional Application No. 60/______). The entire teachings of theabove applications are incorporated herein by reference.

BACKGROUND OF THE INVENTION

[0002] Osteoarthritis (OA) is a slowly progressive, irreversible, oftenmonoarticular disease characterized by pain and loss of function (Mankinand Brandt, Pathogenesis of Osteoarthritis in “Textbook ofRheumatology”, Kelly, et al., (eds.) 3rd edition, W. B. Saunders Co.,Philadelphia, pp.14699-111471 (1989)) and Dean, Arth. Rheum. 20 (Suppl.2):2 (1991)). The underlying cause of the pain and debilitation is thecartilage degradation that occurs as a result of the disease. A typicalend-stage clinical picture includes complete erosion of theweight-bearing articular cartilage, requiring total joint replacement.There is no therapeutic approach available that will slow the clinicalprogression of osteoarthritis, although steroids and non-steroidalanti-inflammatory drugs are used to ameliorate the pain and inflammationassociated with the disease.

SUMMARY OF THE INVENTION

[0003] The present invention relates to isolated nucleic acid moleculesassociated by linkage studies to osteoarthritis (referred to herein as“a variant MATN3 nucleic acid”) and encoding protein associated withosteoarthritis. In one embodiment, the isolated nucleic acid moleculecomprises a nucleotide sequence selected from the group consisting ofSEQ ID NO: 1 and comprising at least one polymorphism selected fromthose set forth in Table 3 or FIGS. 5A-5C and the complements thereof.In a preferred embodiment, the polymorphism is present at nucleotide47928 of SEQ ID NO: 1 (T or C allele).

[0004] The invention further relates to a nucleic acid molecule whichhybridizes under high stringency conditions to a nucleotide sequenceselected from the group consisting of SEQ ID NO: 1 and comprising atleast one polymorphism selected from those set forth in Table 3 or FIGS.5A-5C and the complements thereof.

[0005] The invention further provides a method for assaying the presenceof a nucleic acid molecule comprising all or a portion of the gene in asample, comprising contacting said sample with a second nucleic acidmolecule comprising a nucleotide sequence selected from the groupconsisting of SEQ ID NO: 1 and comprising at least one polymorphismselected from those set forth in Table 3 or FIGS. 5A-5C and thecomplements thereof under conditions appropriate for selectivehybridization.

[0006] The invention also relates to a vector comprising an isolatednucleic acid molecule selected from the group consisting of SEQ ID NO: 1and comprising at least one polymorphism selected from those set forthin Table 3 or FIGS. 5A-5C, operatively linked to a regulatory sequence,as well as to a recombinant host cell comprising the vector. Theinvention also provides a method for preparing a polypeptide encoded byan isolated nucleic acid molecule of the invention selected from thegroup consisting of SEQ ID NO: 1 and comprising at least onepolymorphism selected from those set forth in Table 3 or FIGS. 5A-5C,comprising culturing a recombinant host cell of the invention underconditions suitable for expression of said nucleic acid molecule.

[0007] The invention further provides an isolated polypeptide encoded byisolated nucleic acid molecules of the invention. In a particularembodiment, the polypeptide comprises the amino acid sequence of SEQ IDNO: 2, 3, 4, 5, 6, 7, 8 or 9. The invention also relates to an isolatedpolypeptide comprising an amino acid sequence which is greater thanabout 90 or 95 percent identical to the amino acid sequence of SEQ IDNO: 2, 3, 4, 5, 6, 7, 8 or 9.

[0008] The invention also relates to an antibody, or an antigen-bindingfragment thereof, which selectively binds to a polypeptide of theinvention, as well as to a method for assaying the presence of apolypeptide encoded by an isolated nucleic acid molecule of theinvention in a sample, comprising contacting said sample with anantibody which specifically binds to the encoded polypeptide.

[0009] The invention further relates to methods of diagnosing apredisposition to osteoarthritis. The methods of diagnosing apredisposition to osteoarthritis in an individual include detecting thepresence of an alteration in the MATN3 gene, as well as detectingalterations in expression of a MATN3 polypeptide. The alterations inexpression can be quantitative, qualitative, or both quantitative andqualitative.

[0010] The invention additionally relates to an assay for identifyingagents which alter (e.g., enhance or inhibit) the activity or expressionof a MATN3 polypeptide. For example, a cell, cellular fraction, orsolution containing MATN3 polypeptide or an active fragment orderivative thereof, can be contacted with an agent to be tested, and thelevel of MATN3 polypeptide expression or activity can be assessed.Agents that enhance or inhibit MATN3 polypeptide expression or activityare also included in the current invention, as are methods of altering(enhancing or inhibiting) MATN3 polypeptide expression or activity bycontacting a cell containing MATN3 and/or polypeptide, or by contactingthe MATN3 polypeptide, with an agent that enhances or inhibitsexpression or activity of MATN3 or polypeptide.

[0011] Additionally, the invention pertains to pharmaceuticalcompositions comprising the nucleic acids of the invention, thepolypeptides of the invention, and/or the agents that alter activity ofa MATN3 polypeptide. The invention further pertains to methods oftreating osteoarthritis, by administering MATN3 therapeutic agents, suchas nucleic acids of the invention, polypeptides of the invention, theagents that alter activity of a MATN3 polypeptide, or compositionscomprising the nucleic acids, polypeptides, and/or the agents that alteractivity of a MATN3 polypeptide.

[0012] The invention further provides a method of diagnosingsusceptibility to osteoarthritis in an individual comprising screeningfor an at-risk haplotype in the MATN3 gene that is more frequentlypresent in an individual susceptible to osteoarthritis, compared to thefrequency of its presence in a healthy individual, wherein the presenceof the at-risk haplotype is indicative of a susceptibility toosteoarthritis.

[0013] The invention also relates to a method of diagnosingsusceptibility to osteoarthritis in an individual, comprising screeningfor an at-risk haplotype in the matrilin-3 gene that is more frequentlypresent in an individual susceptible to osteoarthritis (affected),compared to the frequency of its presence in a healthy individual(control), wherein the presence of the at-risk haplotype is indicativeof a susceptibility to osteoarthritis. In one embodiment, the at-riskhaplotype is characterized by the presence of at least one polymorphismat nucleic acid positions 58162, 57927, 56822, 47929, 45434, 45317,45178 and 45010, relative to SEQ ID NO: 1.

[0014] The invention further relates to a kit for diagnosingsusceptibility to osteoarthritis in an individual comprising: primersfor nucleic acid amplification of a region of the matrilin-3 genecomprising an at-risk haplotype, wherein the primers comprise a segmentof nucleic acids of length suitable for nucleic acid amplification,selected from the group consisting of a polymorphism at nucleic acidposition 58162, 57927, 56822, 47929, 45434, 45317, 45178 and 45010,relative to SEQ ID NO: 1 and combinations thereof.

BRIEF DESCRIPTION OF THE DRAWINGS

[0015] The foregoing and other objects, features and advantages of theinvention will be apparent from the following more particulardescription of preferred embodiments of the invention, as illustrated inthe accompanying drawings.

[0016]FIGS. 1.1 to 1.35 show the nucleic acid sequence containing andsurrounding the gene for human MATN3 (SEQ ID NO: 1). Coding sequences ofthe exons are underlined.

[0017]FIGS. 2A to 2C show the exon/intron boundaries of the gene forhuman MATN3 (SEQ ID NOs: 2-9 (amino acid) and SEQ ID NOs: 13-20(nucleotide)). Characterized polymorphisms are labeled above the aminoacids. The coding region is shown within the brackets. Knownpolymorphisms are indicated by asterisks.

[0018]FIG. 3 shows an alignment of amino acid residues for all 4 EGFdomains from MATN3 from human (HuEGF1 to 4) (SEQ ID NOs: 10, 11, 12 and43), mouse (MouEGF1 to 4) (SEQ ID NOs: 44, 45, 46 and 47) and chicken(ChEGF1 to 4) (SEQ ID NOs: 48, 49, 50 and 51). Residues conserved in allEGF domains are indicated by an asterisk. The predicted missensemutation at amino acid position 303 in MATN3 from human protein sequencechanges the threonine-residue (boldface) to methionine. GenBankAccession Numbers for matrilin-3 protein sequences used in alignmentsherein are AJ224741 (human), Y10521 (mouse) and AJ000055 (chicken).

[0019]FIG. 4 is a pedigree showing patients with the Thr/Met mutation atposition 47928. The left side of the symbol filled indicates thumb OA.The right side of the symbol filled indicates finger OA.

[0020]FIGS. 5A to 5C show inserts/deletions occurring at specificpositions of the matrilin-3 gene, as indicated, which were found bysequencing genomic DNA.

[0021]FIGS. 6A to 6B shows the nucleotide sequences of primers used forPCR amplification of DNA sequences of the MATN3 gene.

DETAILED DESCRIPTION OF THE INVENTION

[0022] As described herein, Applicant has completed a genome wide scanon patients with two main subsets of osteoarthritis of the hand; fingerand thumb. The MATN3 gene on chromosome 2 has been identified throughlinkage studies to be associated with osteoarthritis. Until now therehave been no known linkage studies of osteoarthritis in humans showingsignificant connection to this region of the chromosome. Based on thelinkage studies conducted, Applicant has discovered a directrelationship between the MATN3 gene and osteoarthritis. Although theMATN3 gene from normal individuals is known, there have been no studiesdirectly investigating MATN3 and osteoarthritis. Moreover, there havebeen no variant forms reported that have been associated withosteoarthritis. The full sequence of the MATN3 gene (normal gene) isshown in FIGS. 1.1 to 1.35 and SEQ ID NO: 1. It should be understoodthat the nucleic acids and their gene products embraced by the inventioninclude the nucleotide sequence set forth in FIGS. 1.1 to 1.35 andfurther comprise at least one polymorphism as shown in Table 3 or FIGS.5A-5C, and may optionally comprise at least one polymorphism shown inFIGS. 2A to 2C.

[0023] Nucleic Acids of the Invention

[0024] MATN3 Nucleic Acids, Portions and Variants

[0025] Accordingly, the invention pertains to an isolated nucleic acidmolecule comprising a variant form of the human MATN3 gene. The term,“variant MATN3”, as used herein, refers to an isolated nucleic acidmolecule on chromosome 2 having at least one altered nucleotide that isassociated with a susceptibility to osteoarthritis, and also to aportion or fragment of the isolated nucleic acid molecule (e.g., cDNA orthe gene) that encodes MATN3 polypeptide (e.g., the polypeptide havingSEQ ID NO: 2, 3, 4, 5, 6, 7, 8 or 9, as shown in FIGS. 2A to 2C andoptionally comprising at least one polymorphism (e.g., a SNP, aninsertion, or a deletion of one or more nucleotides) as set forth inTable 3 or FIGS. 5A-5C, or another splicing variant of a MATN3polypeptide). In one embodiment, the isolated nucleic acid moleculecomprises the sequence of SEQ ID NO: 1 or the complement of SEQ ID NO:1, except that one or more nucleotide polymorphisms as shown in Table 3or FIGS. 5A-5C are also present. In a particularly preferred embodiment,the isolated nucleic acid molecules comprises at least one of exons 1-8(SEQ ID NOs: 13-20) as shown in FIGS. 2A to 2C.

[0026] The isolated nucleic acid molecules of the present invention canbe RNA, for example, mRNA, or DNA, such as cDNA and genomic DNA. DNAmolecules can be double-stranded or single-stranded; single stranded RNAor DNA can be either the coding, or sense, strand or the non-coding, orantisense, strand. The nucleic acid molecule can include all or aportion of the coding sequence of the gene and can further compriseadditional non-coding sequences such as introns and non-coding 3′ and 5′sequences (including regulatory sequences, for example). Additionally,the nucleic acid molecule can be fused to a marker sequence, forexample, a sequence that encodes a polypeptide to assist in isolation orpurification of the polypeptide. Such sequences include, but are notlimited to, those which encode a glutathione-S-transferase (GST) fusionprotein and those which encode a hemagglutinin A (HA) polypeptide markerfrom influenza.

[0027] An “isolated” nucleic acid molecule, as used herein, is one thatis separated from nucleic acids which normally flank the gene ornucleotide sequence (as in genomic sequences) and/or has been completelyor partially purified from other transcribed sequences (e.g., as in anRNA library). For example, an isolated nucleic acid of the invention maybe substantially isolated with respect to the complex cellular milieu inwhich it naturally occurs, or culture medium when produced byrecombinant techniques, or chemical precursors or other chemicals whenchemically synthesized. In some instances, the isolated material willform part of a composition (for example, a crude extract containingother substances), buffer system or reagent mix. In other circumstances,the material may be purified to essential homogeneity, for example, asdetermined by PAGE or column chromatography such as HPLC. Preferably, anisolated nucleic acid molecule comprises at least about 50, 80, 90 or95% (on a molar basis) of all macromolecular species present. Withregard to genomic DNA, the term “isolated” also can refer to nucleicacid molecules which are separated from the chromosome with which thegenomic DNA is naturally associated. For example, the isolated nucleicacid molecule can contain less than about 5 kb, 4 kb, 3 kb, 2 kb, 1 kb,0.5 kb or 0.1 kb of nucleotides which flank the nucleic acid molecule inthe genomic DNA of the cell from which the nucleic acid molecule isderived.

[0028] The nucleic acid molecule can be fused to other coding orregulatory sequences and still be considered isolated. Thus, recombinantDNA contained in a vector is included in the definition of “isolated” asused herein. Also, isolated nucleic acid molecules include recombinantDNA molecules in heterologous host cells, as well as partially orsubstantially purified DNA molecules in solution. “Isolated” nucleicacid molecules also encompass in vivo and in vitro RNA transcripts ofthe DNA molecules of the present invention. An isolated nucleic acidmolecule or nucleotide sequence can include a nucleic acid molecule ornucleotide sequence which is synthesized chemically or by recombinantmeans. Therefore, recombinant DNA contained in a vector are included inthe definition of “isolated” as used herein. Also, isolated nucleotidesequences include recombinant DNA molecules in heterologous organisms,as well as partially or substantially purified DNA molecules insolution. In vivo and in vitro RNA transcripts of the DNA molecules ofthe present invention are also encompassed by “isolated” nucleotidesequences. Such isolated nucleotide sequences are useful in themanufacture of the encoded polypeptide, as probes for isolatinghomologous sequences (e.g., from other mammalian species), for genemapping (e.g., by in situ hybridization with chromosomes), or fordetecting expression of the gene in tissue (e.g., human tissue), such asby Northern blot analysis.

[0029] The present invention also pertains to variant nucleic acidmolecules which are not necessarily found in nature but which encode aMATN3 polypeptide (e.g., a polypeptide having the amino acid sequence ofSEQ ID NO: 2, 3, 4, 5, 6, 7, 8, or 9), or another splicing variant ofMATN3 polypeptide or polymorphic variant thereof. Thus, for example, DNAmolecules which comprise a sequence that is different from thenaturally-occurring nucleotide sequence but which, due to the degeneracyof the genetic code, encode a MATN3 polypeptide of the present inventionare also the subject of this invention. The invention also encompassesnucleotide sequences encoding portions (fragments), or encoding variantpolypeptides such as analogues or derivatives of the MATN3 polypeptide.Such variants can be naturally-occurring, such as in the case of allelicvariation or single nucleotide polymorphisms, ornon-naturally-occurring, such as those induced by various mutagens andmutagenic processes. Intended variations include, but are not limitedto, addition, deletion and substitution of one or more nucleotides whichcan result in conservative or non-conservative amino acid changes,including additions and deletions. Preferably the nucleotide (and/orresultant amino acid) changes are silent or conserved; that is, they donot alter the characteristics or activity of the MATN3 polypeptide. Inone preferred embodiment, the nucleotide sequences are fragments thatcomprise one or more polymorphic microsatellite markers. In anotherpreferred embodiment, the nucleotide sequences are fragments thatcomprise one or more single nucleotide polymorphisms in the MATN3 gene.

[0030] Other alterations of the nucleic acid molecules of the inventioncan include, for example, labeling, methylation, intemucleotidemodifications such as uncharged linkages (e.g., methyl phosphonates,phosphotriesters, phosphoamidates, carbamates), charged linkages (e.g.,phosphorothioates, phosphorodithioates), pendent moieties (e.g.,polypeptides), intercalators (e.g., acridine, psoralen), chelators,alkylators, and modified linkages (e.g., alpha anomeric nucleic acids).Also included are synthetic molecules that mimic nucleic acid moleculesin the ability to bind to a designated sequences via hydrogen bondingand other chemical interactions. Such molecules include, for example,those in which peptide linkages substitute for phosphate linkages in thebackbone of the molecule.

[0031] The invention also pertains to nucleic acid molecules whichhybridize under high stringency hybridization conditions, such as forselective hybridization, to a nucleotide sequence described herein(e.g., nucleic acid molecules which specifically hybridize to anucleotide sequence encoding polypeptides described herein, and,optionally, have an activity of the polypeptide). In one embodiment, theinvention includes variants described herein which hybridize under highstringency hybridization conditions (e.g., for selective hybridization)to a nucleotide sequence comprising a nucleotide sequence selected fromSEQ ID NO: 1 and comprising at least one polymorphism as shown in Table3 or FIGS. 5A-5C or the complements thereof. In another embodiment, theinvention includes variants described herein which hybridize under highstringency hybridization conditions (e.g., for selective hybridization)to a nucleotide sequence encoding an amino acid sequence selected fromSEQ ID NO: 2, 3, 4, 5, 6, 7, 8 or 9 or polymorphic variant thereof. In apreferred embodiment, the variant which hybridizes under high stringencyhybridizations has an activity of MATN3. Such nucleic acid molecules canbe detected and/or isolated by specific hybridization (e.g., under highstringency conditions). “Specific hybridization,” as used herein, refersto the ability of a first nucleic acid to hybridize to a second nucleicacid in a manner such that the first nucleic acid does not hybridize toany nucleic acid other than to the second nucleic acid (e.g., when thefirst nucleic acid has a higher similarity to the second nucleic acidthan to any other nucleic acid in a sample wherein the hybridization isto be performed). “Stringency conditions” for hybridization is a term ofart which refers to the incubation and wash conditions, e.g., conditionsof temperature and buffer concentration, which permit hybridization of aparticular nucleic acid to a second nucleic acid; the first nucleic acidmay be perfectly (i.e., 100%) complementary to the second, or the firstand second may share some degree of complementarity which is less thanperfect (e.g., 70%, 75%, 85%, 90%, 95%). For example, certain highstringency conditions can be used which distinguish perfectlycomplementary nucleic acids from those of less complementarity. “Highstringency conditions”, “moderate stringency conditions” and “lowstringency conditions” for nucleic acid hybridizations are explained onpages 2.10.1-2.10.16 and pages 6.3.1-6.3.6 in Current Protocols inMolecular Biology (Ausubel, F. M. et al., “Current Protocols inMolecular Biology”, John Wiley & Sons, (1998)), the entire teachings ofwhich are incorporated by reference herein). The exact conditions whichdetermine the stringency of hybridization depend not only on ionicstrength (e.g., 0.2×SSC, 0.1×SSC), temperature (e.g., room temperature,42° C., 68° C.) and the concentration of destabilizing agents such asformamide or denaturing agents such as SDS, but also on factors such asthe length of the nucleic acid sequence, base composition, percentmismatch between hybridizing sequences and the frequency of occurrenceof subsets of that sequence within other non-identical sequences. Thus,equivalent conditions can be determined by varying one or more of theseparameters while maintaining a similar degree of identity or similaritybetween the two nucleic acid molecules. Typically, conditions are usedsuch that sequences at least about 60%, at least about 70%, at leastabout 80%, at least about 90% or at least about 95% or more identical toeach other remain hybridized to one another. By varying hybridizationconditions from a level of stringency at which no hybridization occursto a level at which hybridization is first observed, conditions whichwill allow a given sequence to hybridize (e.g., selectively) with themost similar sequences in the sample can be determined.

[0032] Exemplary conditions are described in Krause, M. H. and S. A.Aaronson (1991) Methods in Enzymology, 200:546-556. Also, in, Ausubel,et al., “Current Protocols in Molecular Biology”, John Wiley & Sons,(1998), which describes the determination of washing conditions formoderate or low stringency conditions. Washing is the step in whichconditions are usually set so as to determine a minimum level ofcomplementarity of the hybrids. Generally, starting from the lowesttemperature at which only homologous hybridization occurs, each ° C. bywhich the final wash temperature is reduced (holding SSC concentrationconstant) allows an increase by 1% in the maximum extent of mismatchingamong the sequences that hybridize. Generally, doubling theconcentration of SSC results in an increase in T_(m) of ˜17° C. Usingthese guidelines, the washing temperature can be determined empiricallyfor high, moderate or low stringency, depending on the level of mismatchsought.

[0033] For example, a low stringency wash can comprise washing in asolution containing 0.2×SSC/0.1% SDS for 10 min at room temperature; amoderate stringency wash can comprise washing in a prewarmed solution(42° C.) solution containing 0.2×SSC/0.1% SDS for 15 min at 42° C.; anda high stringency wash can comprise washing in prewarmed (68° C.)solution containing 0.1×SSC/0.1%SDS for 15 min at 68° C. Furthermore,washes can be performed repeatedly or sequentially to obtain a desiredresult as known in the art. Equivalent conditions can be determined byvarying one or more of the parameters given as an example, as known inthe art, while maintaining a similar degree of identity or similaritybetween the target nucleic acid molecule and the primer or probe used.

[0034] The percent homology or identity of two nucleotide or amino acidsequences can be determined by aligning the sequences for optimalcomparison purposes (e.g., gaps can be introduced in the sequence of afirst sequence for optimal alignment). The nucleotides or amino acids atcorresponding positions are then compared, and the percent identitybetween the two sequences is a function of the number of identicalpositions shared by the sequences (i.e., % identity=# of identicalpositions/total # of positions×100). When a position in one sequence isoccupied by the same nucleotide or amino acid residue as thecorresponding position in the other sequence, then the molecules arehomologous at that position. As used herein, nucleic acid or amino acid“homology” is equivalent to nucleic acid or amino acid “identity”. Incertain embodiments, the length of a sequence aligned for comparisonpurposes is at least 30%, for example, at least 40%, in certainembodiments at least 60%, and in other embodiments at least 70%, 80%,90% or 95% of the length of the reference sequence. The actualcomparison of the two sequences can be accomplished by well-knownmethods, for example, using a mathematical algorithm. A preferred,non-limiting example of such a mathematical algorithm is described inKarlin et al. (1993) Proc. Natl. Acad. Sci. USA 90:5873-5877. Such analgorithm is incorporated into the NBLAST and XBLAST programs (version2.0) as described in Altschul et al. (1997) Nucleic Acids Res.25:389-3402. When utilizing BLAST and Gapped BLAST programs, the defaultparameters of the respective programs (e.g., NBLAST) can be used. In oneembodiment, parameters for sequence comparison can be set at score=100,wordlength=12, or can be varied (e.g., W=5 or W=20).

[0035] Another preferred, non-limiting example of a mathematicalalgorithm utilized for the comparison of sequences is the algorithm ofMyers and Miller, CABIOS (1989). Such an algorithm is incorporated intothe ALIGN program (version 2.0) which is part of the GCG sequencealignment software package. When utilizing the ALIGN program forcomparing amino acid sequences, a PAM120 weight residue table, a gaplength penalty of 12, and a gap penalty of 4 can be used. Additionalalgorithms for sequence analysis are known in the art and includeADVANCE and ADAM as described in Torellis and Robotti (1994) Comput.Appl. Biosci., 10:3-5; and FASTA described in Pearson and Lipman (1988)Proc. Natl. Acad. Sci. USA 85:2444-2448.

[0036] In another embodiment, the percent identity between two aminoacid sequences can be accomplished using the GAP program in the GCGsoftware package (available from Accelrys, San Diego, Calif.) usingeither a Blossom 63 matrix or a PAM250 matrix, and a gap weight of 12,10, 8, 6, or 4 and a length weight of 2, 3, or 4. In yet anotherembodiment, the percent identity between two nucleic acid sequences canbe accomplished using the GAP program in the GCG software package, usinga gap weight of 50 and a length weight of 3.

[0037] The present invention also provides isolated nucleic acidmolecules that contain a fragment or portion that hybridizes underhighly stringent conditions to a nucleotide sequence comprising anucleotide sequence selected from SEQ ID NO: 1 and comprising at leastone polymorphism as shown in Table 3 or FIGS. 5A-5C and the complementsthereof, and also provides isolated nucleic acid molecules that containa fragment or portion that hybridizes under highly stringent conditionsto a nucleotide sequence encoding an amino acid sequence selected fromSEQ ID NO: 2, 3, 4, 5, 6, 7, 8 or 9, or a polymorphic variant thereof.The nucleic acid fragments of the invention are at least about 15,preferably at least about 18, 20, 23 or 25 nucleotides, and can be 30,40, 50, 100, 200 or more nucleotides in length. Longer fragments, forexample, 30 or more nucleotides in length, which encode antigenicpolypeptides described herein are particularly useful, such as for thegeneration of antibodies as described below.

[0038] Probes and Primers

[0039] In a related aspect, the nucleic acid fragments of the inventionare used as probes or primers in assays such as those described herein.“Probes” or “primers” are oligonucleotides that hybridize in abase-specific manner to a complementary strand of nucleic acidmolecules. Such probes and primers include polypeptide nucleic acids, asdescribed in Nielsen et al. (1991) Science, 254:1497-1500.

[0040] A probe or primer comprises a region of nucleotide sequence thathybridizes to at least about 15, for example about 20-25, and in certainembodiments about 40, 50 or 75, consecutive nucleotides of a nucleicacid molecule comprising a contiguous nucleotide sequence selected fromthe group consisting of SEQ ID NOs: 1 and 13-20 or a polymorphic variantthereof. In other embodiments, a probe or primer comprises 100 or fewernucleotides, in certain embodiments from 6 to 50 nucleotides, forexample from 12 to 30 nucleotides. In other embodiments, the probe orprimer is at least 70% identical to the contiguous nucleotide sequenceor to the complement of the contiguous nucleotide sequence, for exampleat least 80% identical, in certain embodiments at least 90% identical,and in other embodiments at least 95% identical, or even capable ofselectively hybridizing to the contiguous nucleotide sequence or to thecomplement of the contiguous nucleotide sequence. Often, the probe orprimer further comprises a label, e.g., radioisotope, fluorescentcompound, enzyme, or enzyme co-factor.

[0041] The nucleic acid molecules of the invention such as thosedescribed above can be identified and isolated using standard molecularbiology techniques and the sequence information provided herein. Forexample, nucleic acid molecules can be amplified and isolated by thepolymerase chain reaction using synthetic oligonucleotide primersdesigned based on one or more of the sequences provided in SEQ ID NO: 1and comprising at least one polymorphism shown in Table 3 or FIGS.5A-5C, and/or the complements thereof, or designed based on nucleotidesbased on sequences encoding one or more of the amino acid sequencesprovided herein. See generally PCR Technology: Principles andApplications for DNA Amplification (ed. H. A. Erlich, Freeman Press, NY,N.Y., 1992); PCR Protocols: A Guide to Methods and Applications (Eds.Innis, et al., Academic Press, San Diego, Calif., 1990); Mattila et al.(1991) Nucleic Acids Res., 19:4967; Eckert et al. (1991) PCR Methods andApplications, 1:17; PCR (eds. McPherson et al., IRL Press, Oxford); andU.S. Pat. No. 4,683,202. The nucleic acid molecules can be amplifiedusing cDNA, mRNA or genomic DNA as a template, cloned into anappropriate vector and characterized by DNA sequence analysis.

[0042] Other suitable amplification methods include the ligase chainreaction (LCR) (see Wu and Wallace (1989) Genomics 4:560, Landegren etal. (1988) Science 241:1077 , transcription amplification (Kwoh et al.(1989) Proc. Natl. Acad. Sci. USA 86:1173), self-sustained sequencereplication (Guatelli et al. (1990) Proc. Nat. Acad. Sci. USA 87:1874)and nucleic acid based sequence amplification (NASBA). The latter twoamplification methods involve isothermal reactions based on isothermaltranscription, which produce both single stranded RNA (ssRNA) and doublestranded DNA (dsDNA) as the amplification products in a ratio of about30 or 100 to 1, respectively.

[0043] The amplified DNA can be labeled, for example, radiolabeled, andused as a probe for screening a cDNA library derived from human cells,mRNA in zap express, ZIPLOX or other suitable vector. Correspondingclones can be isolated, DNA can obtained following in vivo excision, andthe cloned insert can be sequenced in either or both orientations by artrecognized methods to identify the correct reading frame encoding apolypeptide of the appropriate molecular weight. For example, the directanalysis of the nucleotide sequence of nucleic acid molecules of thepresent invention can be accomplished using well-known methods that arecommercially available. See, for example, Sambrook et al., MolecularCloning, A Laboratory Manual (2nd Ed., CSHP, New York 1989); Zyskind etal., Recombinant DNA Laboratory Manual, (Acad. Press, 1988). Using theseor similar methods, the polypeptide and the DNA encoding the polypeptidecan be isolated, sequenced and further characterized.

[0044] Antisense nucleic acid molecules of the invention can be designedusing the nucleotide sequences of SEQ ID NOs: 13-20 and/or thecomplement of SEQ ID NOs: 13-20, and/or a portion of SEQ ID NOs: 13-20or the complement of a portion of SEQ ID NOs: 13-20 or encoding aportion of SEQ ID NO: 1 wherein the portion of SEQ ID NO: 1 comprises atleast one polymorphism as shown in Table 3 or FIGS. 5A-5C andconstructed using chemical synthesis and enzymatic ligation reactionsusing procedures known in the art. For example, an antisense nucleicacid molecule (e.g., an antisense oligonucleotide) can be chemicallysynthesized using naturally occurring nucleotides or variously modifiednucleotides designed to increase the biological stability of themolecules or to increase the physical stability of the duplex formedbetween the antisense and sense nucleic acids, e.g., phosphorothioatederivatives and acridine substituted nucleotides can be used.Alternatively, the antisense nucleic acid molecule can be producedbiologically using an expression vector into which a nucleic acidmolecule has been subcloned in an antisense orientation (i.e., RNAtranscribed from the inserted nucleic acid molecule will be of anantisense orientation to a target nucleic acid of interest).

[0045] In general, the isolated nucleic acid sequences of the inventioncan be used as molecular weight markers on Southern gels, and aschromosome markers which are labeled to map related gene positions. Thenucleic acid sequences can also be used to compare with endogenous DNAsequences in patients to identify genetic disorders (e.g., apredisposition for or susceptibility to osteoarthritis), and as probes,such as to hybridize and discover related DNA sequences or to subtractout known sequences from a sample. The nucleic acid sequences can befurther used to derive primers for genetic fingerprinting, to raiseanti-polypeptide antibodies using DNA immunization techniques, and as anantigen to raise anti-DNA antibodies or elicit immune responses.Portions or fragments of the nucleotide sequences identified herein (andthe corresponding complete gene sequences) can be used in numerous waysas polynucleotide reagents. For example, these sequences can be used to:(i) map their respective genes on a chromosome; and, thus, locate generegions associated with genetic disease; (ii) identify an individualfrom a minute biological sample (tissue typing); and (iii) aid inforensic identification of a biological sample. Additionally, thenucleotide sequences of the invention can be used to identify andexpress recombinant polypeptides for analysis, characterization ortherapeutic use, or as markers for tissues in which the correspondingpolypeptide is expressed, either constitutively, during tissuedifferentiation, or in diseased states. The nucleic acid sequences canadditionally be used as reagents in the screening and/or diagnosticassays described herein, and can also be included as components of kits(e.g., reagent kits) for use in the screening and/or diagnostic assaysdescribed herein.

[0046] Vectors

[0047] Another aspect of the invention pertains to nucleic acidconstructs comprising a nucleic acid molecule of SEQ ID NO: 1 andcomprising at least one polymorphism shown in Table 3 or FIGS. 5A-5C andthe complements thereof (or a portion thereof). Yet another aspect ofthe invention pertains to nucleic acid constructs containing a nucleicacid molecule encoding the amino acid sequence of SEQ ID NO: 2-9 orpolymorphic variants thereof. The constructs comprise a vector (e.g., anexpression vector) into which a sequence of the invention has beeninserted in a sense or antisense orientation. As used herein, the term“vector” refers to a nucleic acid molecule capable of transportinganother nucleic acid to which it has been linked. One type of vector isa “plasmid”, which refers to a circular double stranded DNA loop intowhich additional DNA segments can be ligated. Another type of vector isa viral vector, wherein additional DNA segments can be ligated into theviral genome. Certain vectors are capable of autonomous replication in ahost cell into which they are introduced (e.g., bacterial vectors havinga bacterial origin of replication and episomal mammalian vectors). Othervectors (e.g., non-episomal mammalian vectors) are integrated into thegenome of a host cell upon introduction into the host cell, and therebyare replicated along with the host genome. Moreover, certain vectors,expression vectors, are capable of directing the expression of genes towhich they are operably linked. In general, expression vectors ofutility in recombinant DNA techniques are often in the form of plasmids.However, the invention is intended to include such other forms ofexpression vectors, such as viral vectors (e.g., replication defectiveretroviruses, adenoviruses and adeno-associated viruses) that serveequivalent functions.

[0048] Preferred recombinant expression vectors of the inventioncomprise a nucleic acid molecule of the invention in a form suitable forexpression of the nucleic acid molecule in a host cell. This means thatthe recombinant expression vectors include one or more regulatorysequences, selected on the basis of the host cells to be used forexpression, which is operably linked to the nucleic acid sequence to beexpressed. Within a recombinant expression vector, “operably oroperatively linked” is intended to mean that the nucleotide sequence ofinterest is linked to the regulatory sequence(s) in a manner whichallows for expression of the nucleotide sequence (e.g., in an in vitrotranscription/translation system or in a host cell when the vector isintroduced into the host cell). The term “regulatory sequence” isintended to include promoters, enhancers and other expression controlelements (e.g., polyadenylation signals). Such regulatory sequences aredescribed, for example, in Goeddel, Gene Expression Technology: Methodsin Enzymology 185, Academic Press, San Diego, Calif. (1990). Regulatorysequences include those which direct constitutive expression of anucleotide sequence in many types of host cell and those which directexpression of the nucleotide sequence only in certain host cells (e.g.,tissue-specific regulatory sequences). It will be appreciated by thoseskilled in the art that the design of the expression vector can dependon such factors as the choice of the host cell to be transformed and thelevel of expression of polypeptide desired. The expression vectors ofthe invention can be introduced into host cells to thereby producepolypeptides, including fusion polypeptides, encoded by nucleic acidmolecules as described herein.

[0049] The recombinant expression vectors of the invention can bedesigned for expression of a polypeptide of the invention in prokaryoticor eukaryotic cells, e.g., bacterial cells such as E. coli, insect cells(using baculovirus expression vectors), yeast cells or mammalian cells.Suitable host cells are discussed further in Goeddel (Gene ExpressionTechnology: Methods in Enzymology 185, Academic Press, San Diego, Calif.(1990)). Alternatively, the recombinant expression vector can betranscribed and translated in vitro, for example, using T7 promoterregulatory sequences and T7 polymerase.

[0050] Another aspect of the invention pertains to host cells into whicha recombinant expression vector of the invention has been introduced.The terms “host cell” and “recombinant host cell” are usedinterchangeably herein. It is understood that such terms refer not onlyto the particular subject cell but also to the progeny or potentialprogeny of such a cell. Because certain modifications may occur insucceeding generations due to either mutation or environmentalinfluences, such progeny may not, in fact, be identical to the parentcell, but are still included within the scope of the term as usedherein.

[0051] A host cell can be any prokaryotic or eukaryotic cell. Forexample, a nucleic acid molecule of the invention can be expressed inbacterial cells (e.g., E. coli), insect cells, yeast or mammalian cells(such as Chinese hamster ovary cells (CHO) or COS cells). Other suitablehost cells are known to those skilled in the art.

[0052] Vector DNA can be introduced into prokaryotic or eukaryotic cellsvia conventional transformation or transfection techniques. As usedherein, the terms “transformation” and “transfection” are intended torefer to a variety of art-recognized techniques for introducing aforeign nucleic acid molecule (e.g., DNA) into a host cell, includingcalcium phosphate or calcium chloride co-precipitation,DEAE-dextran-mediated transfection, lipofection, or electroporation.Suitable methods for transforming or transfecting host cells can befound in Sambrook et al. (Molecular Cloning, A Laboratory Manual (2ndEd., CSHP, New York 1989)), and other laboratory manuals.

[0053] For stable transfection of mammalian cells, it is known that,depending upon the expression vector and transfection technique used,only a small fraction of cells may integrate the foreign DNA into theirgenome. In order to identify and select these integrants, a gene thatencodes a selectable marker (e.g., for resistance to antibiotics) isgenerally introduced into the host cells along with the gene ofinterest. Preferred selectable markers include those that conferresistance to drugs, such as G418, hygromycin and methotrexate. Nucleicacid molecules encoding a selectable marker can be introduced into ahost cell on the same vector as the nucleic acid molecule of theinvention or can be introduced on a separate vector. Cells stablytransfected with the introduced nucleic acid molecule can be identifiedby drug selection (e.g., cells that have incorporated the selectablemarker gene will survive, while the other cells die).

[0054] A host cell of the invention, such as a prokaryotic or eukaryotichost cell in culture, can be used to produce (i.e., express) apolypeptide of the invention. Accordingly, the invention furtherprovides methods for producing a polypeptide using the host cells of theinvention. In one embodiment, the method comprises culturing the hostcell of the invention (into which a recombinant expression vectorencoding a polypeptide of the invention has been introduced) in asuitable medium such that the polypeptide is produced. In anotherembodiment, the method further comprises isolating the polypeptide fromthe medium or the host cell.

[0055] The host cells of the invention can also be used to producenonhuman transgenic animals. For example, in one embodiment, a host cellof the invention is a fertilized oocyte or an embryonic stem cell intowhich a nucleic acid molecule of the invention has been introduced(e.g., an exogenous MATN3 gene, or an exogenous nucleic acid encodingMATN3 polypeptide). Such host cells can then be used to create non-humantransgenic animals in which exogenous nucleotide sequences have beenintroduced into the genome or homologous recombinant animals in whichendogenous nucleotide sequences have been altered. Such animals areuseful for studying the function and/or activity of the nucleotidesequence and polypeptide encoded by the sequence and for identifyingand/or evaluating modulators of their activity. As used herein, a“transgenic animal” is a non-human animal, preferably a mammal, morepreferably a rodent such as a rat or mouse, in which one or more of thecells of the animal includes a transgene. Other examples of transgenicanimals include non-human primates, sheep, dogs, cows, goats, chickensand amphibians. A transgene is exogenous DNA which is integrated intothe genome of a cell from which a transgenic animal develops and whichremains in the genome of the mature animal, thereby directing theexpression of an encoded gene product in one or more cell types ortissues of the transgenic animal. As used herein, an “homologousrecombinant animal” is a non-human animal, preferably a mammal, morepreferably a mouse, in which an endogenous gene has been altered byhomologous recombination between the endogenous gene and an exogenousDNA molecule introduced into a cell of the animal, e.g., an embryoniccell of the animal, prior to development of the animal.

[0056] Methods for generating transgenic animals via embryo manipulationand microinjection, particularly animals such as mice, have becomeconventional in the art and are described, for example, in U.S. Pat.Nos. 4,736,866, 4,870,009 and 4,873,191 and in Hogan, Manipulating theMouse Embryo (Cold Spring Harbor Laboratory Press, Cold Spring Harbor,N.Y., 1986). Methods for constructing homologous recombination vectorsand homologous recombinant animals are described further in Bradley(1991) Current Opinion in Bio/Technology 2:823-829 and in PCTPublication Nos. WO 90/11354, WO 91/01140, WO 92/0968 and WO 93/04169.Clones of the non-human transgenic animals described herein can also beproduced according to the methods described in Wilmut et al. (1997)Nature 385:810-813 and PCT Publication Nos. WO 97/07668 and WO 97/07669.

[0057] Polypeptides of the Invention

[0058] The present invention also pertains to isolated polypeptidesencoded by MATN3 (“MATN3 polypeptides”) and fragments and variantsthereof, as well as polypeptides encoded by nucleotide sequencesdescribed herein (e.g., other splicing variants). The term “polypeptide”refers to a polymer of amino acids, and not to a specific length; thus,peptides, oligopeptides and proteins are included within the definitionof a polypeptide. As used herein, a polypeptide is said to be “isolated”or “purified” when it is substantially free of cellular material when itis isolated from recombinant and non-recombinant cells, or free ofchemical precursors or other chemicals when it is chemicallysynthesized. A polypeptide, however, can be joined to anotherpolypeptide with which it is not normally associated in a cell (e.g., ina “fusion protein”) and still be “isolated” or “purified.”

[0059] The polypeptides of the invention can be purified to homogeneity.It is understood, however, that preparations in which the polypeptide isnot purified to homogeneity are useful. The critical feature is that thepreparation allows for the desired function of the polypeptide, even inthe presence of considerable amounts of other components. Thus, theinvention encompasses various degrees of purity. In one embodiment, thelanguage “substantially free of cellular material” includes preparationsof the polypeptide having less than about 30% (by dry weight) otherproteins (i.e., contaminating protein), less than about 20% otherproteins, less than about 10% other proteins, or less than about 5%other proteins.

[0060] When a polypeptide is recombinantly produced, it can also besubstantially free of culture medium, i.e., culture medium representsless than about 20%, less than about 10%, or less than about 5% of thevolume of the polypeptide preparation. The language “substantially freeof chemical precursors or other chemicals” includes preparations of thepolypeptide in which it is separated from chemical precursors or otherchemicals that are involved in its synthesis. In one embodiment, thelanguage “substantially free of chemical precursors or other chemicals”includes preparations of the polypeptide having less than about 30% (bydry weight) chemical precursors or other chemicals, less than about 20%chemical precursors or other chemicals, less than about 10% chemicalprecursors or other chemicals, or less than about 5% chemical precursorsor other chemicals.

[0061] In one embodiment, a polypeptide of the invention comprises anamino acid sequence encoded by a nucleic acid molecule comprising anucleotide sequence selected from the group consisting of SEQ ID NO: 1comprising at least one polymorphism shown in Table 3 or FIGS. 5A-5C andcomplements and portions thereof, e.g., SEQ ID NOs: 13-20, or a portionor polymorphic variant thereof. However, the polypeptides of theinvention also encompass fragment and sequence variants. Variantsinclude a substantially homologous polypeptide encoded by the samegenetic locus in an organism, i.e., an allelic variant, as well as othersplicing variants. Variants also encompass polypeptides derived fromother genetic loci in an organism, but having substantial homology to apolypeptide encoded by a nucleic acid molecule comprising a nucleotidesequence selected from the group consisting of SEQ ID NO: 1 comprisingat least one polymorphism shown in Table 3 or FIGS. 5A-5C andcomplements and portions thereof, or having substantial homology to apolypeptide encoded by a nucleic acid molecule comprising a nucleotidesequence selected from the group consisting of nucleotide sequencesencoding SEQ ID NO: 1, or polymorphic variants thereof. Variants alsoinclude polypeptides substantially homologous or identical to thesepolypeptides but derived from another organism, i.e., an ortholog.Variants also include polypeptides that are substantially homologous oridentical to these polypeptides that are produced by chemical synthesis.Variants also include polypeptides that are substantially homologous oridentical to these polypeptides that are produced by recombinantmethods.

[0062] As used herein, two polypeptides (or a region of thepolypeptides) are substantially homologous or identical when the aminoacid sequences are at least about 45-55%, in certain embodiments atleast about 70-75%, and in other embodiments at least about 80-85%, andin other embodiments greater than about 90% or more homologous oridentical (e.g., 95%). A substantially homologous amino acid sequence,according to the present invention, will be encoded by a nucleic acidmolecule of the invention, or portion thereof, under stringentconditions as more particularly described above, or will be encoded by anucleic acid molecule hybridizing to a nucleic acid sequence of theinvention, portion thereof or polymorphic variant thereof, understringent conditions as more particularly described thereof.

[0063] The invention also encompasses polypeptides having a lower degreeof identity but having sufficient similarity so as to perform one ormore of the same functions performed by a polypeptide encoded by anucleic acid molecule of the invention. Similarity is determined byconserved amino acid substitution. Such substitutions are those thatsubstitute a given amino acid in a polypeptide by another amino acid oflike characteristics. Conservative substitutions are likely to bephenotypically silent. Typically seen as conservative substitutions arethe replacements, one for another, among the aliphatic amino acids Ala,Val, Leu and Ile; interchange of the hydroxyl residues Ser and Thr,exchange of the acidic residues Asp and Glu, substitution between theamide residues Asn and Gln, exchange of the basic residues Lys and Argand replacements among the aromatic residues Phe and Tyr. Guidanceconcerning which amino acid changes are likely to be phenotypicallysilent are found in Bowie et al. (1990) Science, 247:1306-1310.

[0064] A variant polypeptide can differ in amino acid sequence by one ormore substitutions, deletions, insertions, inversions, fusions, andtruncations or a combination of any of these. Further, variantpolypeptides can be fully functional or can lack function in one or moreactivities. Fully functional variants typically contain onlyconservative variation or variation in non-critical residues or innon-critical regions. Functional variants can also contain substitutionof similar amino acids that result in no change or an insignificantchange in function. Alternatively, such substitutions may positively ornegatively affect function to some degree. Non-functional variantstypically contain one or more non-conservative amino acid substitutions,deletions, insertions, inversions, or truncation or a substitution,insertion, inversion, or deletion in a critical residue or criticalregion.

[0065] Amino acids that are essential for function can be identified bymethods known in the art, such as site-directed mutagenesis oralanine-scanning mutagenesis (Cunningham et al. (1989) Science,244:1081-1085). The latter procedure introduces single alanine mutationsat every residue in the molecule. The resulting mutant molecules arethen tested for biological activity in vitro, or in vitro proliferativeactivity. Sites that are critical for polypeptide activity can also bedetermined by structural analysis such as crystallization, nuclearmagnetic resonance or photoaffinity labeling (Smith et al. (1992) J.Mol. Biol., 224:899-904; de Vos et al. (1992) Science, 255:306-312).

[0066] The invention also includes polypeptide fragments of thepolypeptides of the invention. Fragments can be derived from apolypeptide encoded by a nucleic acid described herein. However, theinvention also encompasses fragments of the variants of the polypeptidesdescribed herein. As used herein, a fragment comprises at least 6contiguous amino acids. Useful fragments include those that retain oneor more of the biological activities of the polypeptide as well asfragments that can be used as an immunogen to generatepolypeptide-specific antibodies.

[0067] Biologically active fragments (peptides which are, for example,6, 9, 12, 15, 16, 20, 30, 35, 36, 37, 38, 39, 40, 50, 100 or more aminoacids in length) can comprise a domain, segment, or motif that has beenidentified by analysis of the polypeptide sequence using well-knownmethods, e.g., signal peptides, extracellular domains, one or moretransmembrane segments or loops, ligand binding regions, zinc fingerdomains, DNA binding domains, acylation sites, glycosylation sites, orphosphorylation sites.

[0068] Fragments can be discrete (not fused to other amino acids orpolypeptides) or can be within a larger polypeptide. Further, severalfragments can be comprised within a single larger polypeptide. In oneembodiment a fragment designed for expression in a host can haveheterologous pre- and pro-polypeptide regions fused to the aminoterminus of the polypeptide fragment and an additional region fused tothe carboxyl terminus of the fragment.

[0069] The invention thus provides chimeric or fusion polypeptides.These comprise a polypeptide of the invention operatively linked to aheterologous protein or polypeptide having an amino acid sequence notsubstantially homologous to the polypeptide. “Operatively linked”indicates that the polypeptide and the heterologous protein are fusedin-frame. The heterologous protein can be fused to the N-terminus orC-terminus of the polypeptide. In one embodiment the fusion polypeptidedoes not affect function of the polypeptide per se. For example, thefusion polypeptide can be a GST-fusion polypeptide in which thepolypeptide sequences are fused to the C-terminus of the GST sequences.Other types of fusion polypeptides include, but are not limited to,enzymatic fusion polypeptides, for example, β-galactosidase fusions,yeast two-hybrid GAL fusions, poly-His fusions and Ig fusions. Suchfusion polypeptides, particularly poly-His fusions, can facilitate thepurification of recombinant polypeptide. In certain host cells (e.g.,mammalian host cells), expression and/or secretion of a polypeptide canbe increased by using a heterologous signal sequence. Therefore, inanother embodiment, the fusion polypeptide contains a heterologoussignal sequence at its N-terminus.

[0070] EP-A-O 464 533 discloses fusion proteins comprising variousportions of immunoglobulin constant regions. The Fc is useful in therapyand diagnosis and thus results, for example, in improved pharmacokineticproperties (EP-A 0232 262). In drug discovery, for example, humanproteins have been fused with Fc portions for the purpose ofhigh-throughput screening assays to identify antagonists. Bennett et al.(1995) Journal of Molecular Recognition 8:52-58 and Johanson et al.(1995) The Journal of Biological Chemistry, 270(16):9459-9471. Thus,this invention also encompasses soluble fusion polypeptides containing apolypeptide of the invention and various portions of the constantregions of heavy or light chains of immunoglobulins of various subclass(IgG, IgM, IgA, IgE).

[0071] A chimeric or fusion polypeptide can be produced by standardrecombinant DNA techniques. For example, DNA fragments coding for thedifferent polypeptide sequences are ligated together in-frame inaccordance with conventional techniques. In another embodiment, thefusion gene can be synthesized by conventional techniques includingautomated DNA synthesizers. Alternatively, PCR amplification of nucleicacid fragments can be carried out using anchor primers which give riseto complementary overhangs between two consecutive nucleic acidfragments which can subsequently be annealed and re-amplified togenerate a chimeric nucleic acid sequence (see Ausubel et al., CurrentProtocols in Molecular Biology, 1992). Moreover, many expression vectorsare commercially available that already encode a fusion moiety (e.g., aGST protein). A nucleic acid molecule encoding a polypeptide of theinvention can be cloned into such an expression vector such that thefusion moiety is linked in-frame to the polypeptide.

[0072] The isolated polypeptide can be purified from cells thatnaturally express it, purified from cells that have been altered toexpress it (recombinant), or synthesized using known protein synthesismethods. In one embodiment, the polypeptide is produced by recombinantDNA techniques. For example, a nucleic acid molecule encoding thepolypeptide is cloned into an expression vector, the expression vectorintroduced into a host cell and the polypeptide expressed in the hostcell. The polypeptide can then be isolated from the cells by anappropriate purification scheme using standard protein purificationtechniques.

[0073] In general, polypeptides of the present invention can be used asa molecular weight marker on SDS-PAGE gels or on molecular sieve gelfiltration columns using art-recognized methods. The polypeptides of thepresent invention can be used to raise antibodies or to elicit an immuneresponse. The polypeptides can also be used as a reagent, e.g., alabeled reagent, in assays to quantitatively determine levels of thepolypeptide or a molecule to which it binds (e.g., a receptor or aligand) in biological fluids. The polypeptides can also be used asmarkers for cells or tissues in which the corresponding polypeptide ispreferentially expressed, either constitutively, during tissuedifferentiation, or in a diseased state. The polypeptides can be used toisolate a corresponding binding agent, e.g., receptor or ligand, suchas, for example, in an interaction trap assay, and to screen for peptideor small molecule antagonists or agonists of the binding interaction.

[0074] Antibodies of the Invention

[0075] Polyclonal and/or monoclonal antibodies that specifically bindone form of the gene product but not to the other form of the geneproduct are also provided. Antibodies are also provided that bind aportion of either the variant or the reference gene product thatcontains the polymorphic site or sites. The invention providesantibodies to the polypeptides and polypeptide fragments of theinvention, e.g., having an amino acid sequence encoded by SEQ ID NO: 2,3, 4, 5, 6, 7, 8 or 9, or a portion thereof, or having an amino acidsequence encoded by a nucleic acid molecule comprising all or a portionof SEQ ID NO: 1 and comprising at least one polymorphism shown in Table3 or FIGS. 5A-5C. The term “antibody” as used herein refers toimmunoglobulin molecules and immunologically active portions ofimmunoglobulin molecules, i.e., molecules that contain an antigenbinding site that specifically binds an antigen. A molecule thatspecifically binds to a polypeptide of the invention is a molecule thatbinds to that polypeptide or a fragment thereof, but does notsubstantially bind other molecules in a sample, e.g., a biologicalsample, which naturally contains the polypeptide. Examples ofimmunologically active portions of immunoglobulin molecules includeF(ab) and F(ab′)₂ fragments which can be generated by treating theantibody with an enzyme such as pepsin. The invention providespolyclonal and monoclonal antibodies that bind to a polypeptide of theinvention. The term “monoclonal antibody” or “monoclonal antibodycomposition”, as used herein, refers to a population of antibodymolecules that contain only one species of an antigen binding sitecapable of immunoreacting with a particular epitope of a polypeptide ofthe invention. A monoclonal antibody composition thus typically displaysa single binding affinity for a particular polypeptide of the inventionwith which it immunoreacts.

[0076] Polyclonal antibodies can be prepared as described above byimmunizing a suitable subject with a desired immunogen, e.g.,polypeptide of the invention or fragment thereof. The antibody titer inthe immunized subject can be monitored over time by standard techniques,such as with an enzyme linked immunosorbent assay (ELISA) usingimmobilized polypeptide. If desired, the antibody molecules directedagainst the polypeptide can be isolated from the mammal (e.g., from theblood) and further purified by well-known techniques, such as protein Achromatography to obtain the IgG fraction. At an appropriate time afterimmunization, e.g., when the antibody titers are highest,antibody-producing cells can be obtained from the subject and used toprepare monoclonal antibodies by standard techniques, such as thehybridoma technique originally described by Kohler and Milstein (1975)Nature 256:495-497, the human B cell hybridoma technique (Kozbor et al.(1983) Immunol. Today 4:72), the EBV-hybridoma technique (Cole et al.,Monoclonal Antibodies and Cancer Therapy, Alan R. Liss, Inc., pp. 77-96(1985)) or trioma techniques. The technology for producing hybridomas iswell known (see generally Current Protocols in Immunology, Coligan etal. (eds.) John Wiley & Sons, Inc., New York, N.Y. (1994)). Briefly, animmortal cell line (typically a myeloma) is fused to lymphocytes(typically splenocytes) from a mammal immunized with an immunogen asdescribed above, and the culture supernatants of the resulting hybridomacells are screened to identify a hybridoma producing a monoclonalantibody that binds a polypeptide of the invention.

[0077] Any of the many well known protocols used for fusing lymphocytesand immortalized cell lines can be applied for the purpose of generatinga monoclonal antibody to a polypeptide of the invention (see, e.g.,Current Protocols in Immunology, Coligan et al. (eds.) John Wiley &Sons, Inc., New York, N.Y. (1994); Galfre et al. (1977) Nature266:55052; R. H. Kenneth, in Monoclonal Antibodies: A New Dimension InBiological Analyses, Plenum Publishing Corp., New York, N.Y. (1980); andLerner (1981) Yale J. Biol. Med. 54:387-402). Moreover, the ordinarilyskilled worker will appreciate that there are many variations of suchmethods that also would be useful.

[0078] Alternative to preparing monoclonal antibody-secretinghybridomas, a monoclonal antibody to a polypeptide of the invention canbe identified and isolated by screening a recombinant combinatorialimmunoglobulin library (e.g., an antibody phage display library) withthe polypeptide to thereby isolate immunoglobulin library members thatbind the polypeptide. Kits for generating and screening phage displaylibraries are commercially available (e.g., the Pharmacia RecombinantPhage Antibody System, Catalog No. 27-9400-01; and the StratageneSurfZAP™ Phage Display Kit, Catalog No. 240612). Additionally, examplesof methods and reagents particularly amenable for use in generating andscreening antibody display library can be found in, for example, U.S.Pat. No. 5,223,409; PCT Publication No. WO 92/18619; PCT Publication No.WO 91/17271; PCT Publication No. WO 92/20791; PCT Publication No. WO92/15679; PCT Publication No. WO 93/01288; PCT Publication No. WO92/01047; PCT Publication No. WO 92/09690; PCT Publication No. WO90/02809; Fuchs et al., (1991) Bio/Technology, 9:1370-1372; Hay et al.,(1992) Hum. Antibod. Hybridomas, 3:81-85; Huse et al.,(1989) Science,246:1275-1281; and Griffiths et al., (1993) EMBO J, 12:725-734.

[0079] Additionally, recombinant antibodies, such as chimeric andhumanized monoclonal antibodies, comprising both human and non-humanportions, which can be made using standard recombinant DNA techniques,are within the scope of the invention. Such chimeric and humanizedmonoclonal antibodies can be produced by recombinant DNA techniquesknown in the art.

[0080] In addition, human antibodies are within the scope of thisinvention. Such human antibodies can be produced, isolated and purifiedby techniques known to one skilled in the art and using standardmethodologies.

[0081] In general, antibodies of the invention (e.g., a monoclonalantibody) can be used to isolate a polypeptide of the invention bystandard techniques, such as affinity chromatography orimmunoprecipitation. A polypeptide-specific antibody can facilitate thepurification of natural polypeptide from cells and of recombinantlyproduced polypeptide expressed in host cells. Moreover, an antibodyspecific for a polypeptide of the invention can be used to detect thepolypeptide (e.g., in a cellular lysate, cell supernatant, or tissuesample) in order to evaluate the abundance and pattern of expression ofthe polypeptide. Antibodies can be used diagnostically to monitorprotein levels in tissue as part of a clinical testing procedure, e.g.,to determine the efficacy of a given treatment regimen. Detection can befacilitated by coupling the antibody to a detectable substance. Examplesof detectable substances include various enzymes, prosthetic groups,fluorescent materials, luminescent materials, bioluminescent materialsand radioactive materials. Examples of suitable enzymes includehorseradish peroxidase, alkaline phosphatase, β-galactosidase, oracetylcholinesterase; examples of suitable prosthetic group complexesinclude streptavidin/biotin and avidin/biotin; examples of suitablefluorescent materials include umbelliferone, fluorescein, fluoresceinisothiocyanate, rhodamine, dichlorotriazinylamine fluorescein, dansylchloride or phycoerythrin; an example of a luminescent material includesluminol; examples of bioluminescent materials include luciferase,luciferin and aequorin, and examples of suitable radioactive materialinclude ¹²⁵I, ¹³¹I, ³⁵S or ³H.

[0082] Diagnostic and Screening Assays of the Invention

[0083] The present invention also pertains to a method of diagnosing oraiding in the diagnosis of osteoarthritis associated with the presenceof a variant form of the MATN3 gene or gene product in an individual.Diagnostic assays can be designed for assessing MATN3 gene expression,or for assessing activity of MATN3 polypeptides of the invention. In oneembodiment, the assays are used in the context of a biological sample(e.g., blood, serum, cells, tissue, synovial fluid) to thereby determinewhether an individual is afflicted with osteoarthritis, or is at riskfor (has a predisposition for or a susceptibility to) developingosteoarthritis. The invention also provides for prognostic (orpredictive) assays for determining whether an individual is susceptibleto developing osteoarthritis. For example, alterations in nucleic acidscan be assayed in a biological sample. Such assays can be used forprognostic or predictive purpose to thereby prophylactically treat anindividual prior to the onset of symptoms associated withosteoarthritis. Another aspect of the invention pertains to assays formonitoring the influence of agents (e.g., drugs, compounds or otheragents) on the expression or activity of polypeptides of the invention,as well as to assays for identifying agents which bind to MATN3polypeptides. These and other assays and agents are described in furtherdetail in the following sections.

[0084] Diagnostic Assays

[0085] The nucleic acids, probes, primers, polypeptides and antibodiesdescribed herein can be used in methods of diagnosis of a susceptibilityto osteoarthritis as well as kits comprising same.

[0086] In one embodiment of the invention, diagnosis of a susceptibilityto osteoarthritis is made by detecting a polymorphism in MATN3 asdescribed herein. The polymorphism can be a change in MATN3, such as theinsertion or deletion of a single nucleotide, or of more than onenucleotide, resulting in a frame shift; the change of at least onenucleotide, resulting in a change in the encoded amino acid; the changeof at least one nucleotide, resulting in the generation of a prematurestop codon; the deletion of several nucleotides, resulting in a deletionof one or more amino acids encoded by the nucleotides; the insertion ofone or several nucleotides, such as by unequal recombination or geneconversion, resulting in an interruption of the coding sequence of thegene; duplication of all or a part of the gene; transposition of all ora part of the gene; or rearrangement of all or a part of the gene. Morethan one such change may be present in a single gene. Such sequencechanges cause a difference in the polypeptide encoded by a MATN3 nucleicacid. For example, if the alteration is a frame shift mutation, theframe shift can result in a change in the encoded amino acids, and/orcan result in the generation of a premature stop codon, causinggeneration of a truncated polypeptide. Alternatively, a polymorphismassociated with a susceptibility to osteoarthritis can be a synonymousalteration in one or more nucleotides (i.e., an alteration that does notresult in a change in the polypeptide encoded by a MATN3 nucleic acid).Such a polymorphism may alter splicing sites, affect the stability ortransport of mRNA, or otherwise affect the transcription or translationof the gene. A MATN3 nucleic acid that has any of the alterationsdescribed above is referred to herein as an “altered nucleic acid.”

[0087] In a first method of diagnosing a susceptibility toosteoarthritis, hybridization methods, such as Southern analysis,Northern analysis, or in situ hybridizations, can be used (see CurrentProtocols in Molecular Biology, Ausubel, F. et al., eds., John Wiley &Sons, including all supplements through 1999). For example, a biologicalsample from a test subject (a “test sample”) of genomic DNA, RNA, orcDNA, is obtained from an individual suspected of having, beingsusceptible to or predisposed for, or carrying a defect for,osteoarthritis (the “test individual”). The individual can be an adult,child, or fetus. The test sample can be from any source which containsgenomic DNA, such as a blood sample, sample of synovial fluid, sample ofamniotic fluid, sample of cerebrospinal fluid, or tissue sample fromskin, muscle, buccal or conjunctival mucosa, placenta, gastrointestinaltract or other organs. A test sample of DNA from fetal cells or tissuecan be obtained by appropriate methods, such as by amniocentesis orchorionic villus sampling. The DNA, RNA, or cDNA sample is then examinedto determine whether a polymorphism in MATN3 is present, and/or todetermine which splicing variant(s) encoded by MATN3 is present. Thepresence of the polymorphism or splicing variant(s) can be indicated byhybridization of the gene in the genomic DNA, RNA, or cDNA to a nucleicacid probe. A “nucleic acid probe”, as used herein, can be a DNA probeor an RNA probe; the nucleic acid probe can contain at least onepolymorphism in MATN3 or contains a nucleic acid encoding a particularsplicing variant of MATN3. The probe can be any of the nucleic acidmolecules described above (e.g., the gene or nucleic acid, a fragment, avector comprising the gene, a probe or primer, etc.).

[0088] To diagnose a susceptibility to osteoarthritis, a hybridizationsample is formed by contacting the test sample containing MATN3, with atleast one nucleic acid probe. A preferred probe for detecting mRNA orgenomic DNA is a labeled nucleic acid probe capable of hybridizing tomRNA or genomic DNA sequences described herein. The nucleic acid probecan be, for example, a full-length nucleic acid molecule, or a portionthereof, such as an oligonucleotide of at least 15, 30, 50, 100, 250 or500 nucleotides in length and sufficient to specifically hybridize understringent conditions to appropriate mRNA or genomic DNA. For example,the nucleic acid probe can be all or a portion of SEQ ID NO: 1 andcomprising at least one polymorphism shown in Table 3 or FIGS. 5A-5C, orthe complements thereof, or a portion thereof; or can be a nucleic acidencoding a portion thereof. Other suitable probes for use in thediagnostic assays of the invention are described above (see e.g., probesand primers discussed under the heading, “Nucleic Acids of theInvention”).

[0089] The hybridization sample is maintained under conditions which aresufficient to allow specific hybridization of the nucleic acid probe toMATN3. “Specific hybridization”, as used herein, indicates exacthybridization (e.g., with no mismatches). Specific hybridization can beperformed under high stringency conditions or moderate stringencyconditions, for example, as described above. In a particularly preferredembodiment, the hybridization conditions for specific hybridization arehigh stringency.

[0090] Specific hybridization, if present, is then detected usingstandard methods. If specific hybridization occurs between the nucleicacid probe and MATN3 in the test sample, then MATN3 has thepolymorphism, or is the splicing variant, that is present in the nucleicacid probe. More than one nucleic acid probe can also be usedconcurrently in this method. Specific hybridization of any one of thenucleic acid probes is indicative of a polymorphism in MATN3, or of thepresence of a particular splicing variant encoding MATN3 and istherefore diagnostic for a susceptibility to osteoarthritis.

[0091] In Northern analysis (see Ausubel, F. et al., eds., “CurrentProtocols in Molecular Biology”, John Wiley & Sons, (1998)) thehybridization methods described above are used to identify the presenceof a polymorphism or a particular splicing variant, associated with asusceptibility to osteoarthritis. For Northern analysis, a test sampleof RNA is obtained from the individual by appropriate means. Specifichybridization of a nucleic acid probe, as described above, to RNA fromthe individual is indicative of a polymorphism in MATN3, or of thepresence of a particular splicing variant encoded by MATN3, and istherefore diagnostic for a susceptibility to osteoarthritis.

[0092] For representative examples of use of nucleic acid probes, see,for example, U.S. Pat. Nos. 5,288,611 and 4,851,330.

[0093] Alternatively, a peptide nucleic acid (PNA) probe can be usedinstead of a nucleic acid probe in the hybridization methods describedabove. PNA is a DNA mimic having a peptide-like, inorganic backbone,such as N-(2-aminoethyl)glycine units, with an organic base (A, G, C, Tor U) attached to the glycine nitrogen via a methylene carbonyl linker(see, for example, Nielsen, P. E. et al. (1994) Bioconjugate Chemistry5, American Chemical Society, p. 1). The PNA probe can be designed tospecifically hybridize to a gene having a polymorphism associated with asusceptibility to osteoarthritis. Hybridization of the PNA probe toMATN3 is diagnostic for a susceptibility to osteoarthritis.

[0094] In another method of the invention, mutation analysis byrestriction digestion can be used to detect an altered nucleic acid orgene, or genes containing a polymorphism(s), if the alteration orpolymorphism in the gene results in the creation or elimination of arestriction site. A test sample containing genomic DNA is obtained fromthe individual. Polymerase chain reaction (PCR) can be used to amplifyMATN3 (and, if necessary, the flanking sequences) in the test sample ofgenomic DNA from the test individual. RFLP analysis is conducted asdescribed (see “Current Protocols in Molecular Biology”, John Wiley &Sons, (1998)). The digestion pattern of the relevant DNA fragmentindicates the presence or absence of the mutation or polymorphism inMATN3, and therefore indicates the presence or absence of thissusceptibility to osteoarthritis.

[0095] Sequence analysis can also be used to detect specificpolymorphisms in MATN3. A test sample of DNA or RNA is obtained from thetest individual. PCR or other appropriate methods can be used to amplifythe gene, and/or its flanking sequences, if desired. The sequence ofMATN3, or a fragment of the gene, or cDNA, or fragment of the cDNA, ormRNA, or fragment of the mRNA, is determined, using standard methods.The sequence of the gene, gene fragment, cDNA, cDNA fragment, mRNA, ormRNA fragment is compared with the known nucleic acid sequence of thegene, cDNA or mRNA, as appropriate. The presence of a polymorphism inMATN3 indicates that the individual has a susceptibility toosteoarthritis.

[0096] Allele-specific oligonucleotides can also be used to detect thepresence of a polymorphism in MATN3, through the use of dot-blothybridization of amplified oligonucleotides with allele-specificoligonucleotide (ASO) probes (see, for example, Saiki et al. (1986)Nature (London) 324:163-166). An “allele-specific oligonucleotide” (alsoreferred to herein as an “allele-specific oligonucleotide probe”) is anoligonucleotide of approximately 10-50 base pairs, preferablyapproximately 15-30 base pairs, that specifically hybridizes to MATN3,and that contains a polymorphism associated with a susceptibility toosteoarthritis. An allele-specific oligonucleotide probe that isspecific for particular polymorphisms in MATN3 can be prepared, usingstandard methods (see Ausubel, F. et al., eds., “Current Protocols inMolecular Biology”, John Wiley & Sons, (1998)). To identifypolymorphisms in the gene that are associated with a susceptibility toosteoarthritis, a test sample of DNA is obtained from the individual.PCR can be used to amplify all or a fragment of MATN3, and its flankingsequences. The DNA containing the amplified MATN3 (or fragment of thegene) is dot-blotted, using standard methods (see Ausubel, F. et al.,eds., “Current Protocols in Molecular Biology”, John Wiley & Sons,(1998)), and the blot is contacted with the oligonucleotide probe. Thepresence of specific hybridization of the probe to the amplified MATN3is then detected. Specific hybridization of an allele-specificoligonucleotide probe to DNA from the individual is indicative of apolymorphism in MATN3, and is therefore indicative of a susceptibilityto osteoarthritis.

[0097] The invention further provides allele-specific oligonucleotidesthat hybridize to the reference or variant allele of a gene or nucleicacid comprising a single nucleotide polymorphism or to the complementthereof. These oligonucleotides can be probes or primers.

[0098] An allele-specific primer hybridizes to a site on target DNAoverlapping a polymorphism and only primes amplification of an allelicform to which the primer exhibits perfect complementarity. See Gibbs(1989) Nucleic Acids Res. 17:2427-2448. This primer is used inconjunction with a second primer, which hybridizes at a distal site.Amplification proceeds from the two primers, resulting in a detectableproduct, which indicates the particular allelic form is present. Acontrol is usually performed with a second pair of primers, one of whichshows a single base mismatch at the polymorphic site and the other ofwhich exhibits perfect complementarity to a distal site. The single-basemismatch prevents amplification and no detectable product is formed. Themethod works best when the mismatch is included in the 3′-most positionof the oligonucleotide aligned with the polymorphism because thisposition is most destabilizing to elongation from the primer (see, e.g.,WO 93/22456).

[0099] In another embodiment, arrays of oligonucleotide probes that arecomplementary to target nucleic acid sequence segments from anindividual, can be used to identify polymorphisms in MATN3. For example,in one embodiment, an oligonucleotide array can be used. Oligonucleotidearrays typically comprise a plurality of different oligonucleotideprobes that are coupled to a surface of a substrate in different knownlocations. These oligonucleotide arrays, also described as“Genechips.TM.,” have been generally described in the art, for example,U.S. Pat. No. 5,143,854 and PCT patent publication Nos. WO 90/15070 and92/10092. These arrays can generally be produced using mechanicalsynthesis methods or light directed synthesis methods which incorporatea combination of photolithographic methods and solid phaseoligonucleotide synthesis methods. See Fodor et al. (1991) Science251:767-777, Pirrung et al., U.S. Pat. No. 5,143,854 (see also PCTApplication No. WO 90/15070) and Fodor et al., PCT Publication No. WO92/10092 and U.S. Pat. No. 5,424,186, the entire teachings of each ofwhich are incorporated by reference herein. Techniques for the synthesisof these arrays using mechanical synthesis methods are described in,e.g., U.S. Pat. No. 5,384,261, the entire teachings of which areincorporated by reference herein. In another example, linear arrays canbe utilized.

[0100] Once an oligonucleotide array is prepared, a nucleic acid ofinterest is hybridized with the array and scanned for polymorphisms.Hybridization and scanning are generally carried out by methodsdescribed herein and also in, e.g., Published PCT Application Nos. WO92/10092 and WO 95/11995 and U.S. Pat. No. 5,424,186, the entireteachings of which are incorporated by reference herein. In brief, atarget nucleic acid sequence which includes one or more previouslyidentified polymorphic markers is amplified by well known amplificationtechniques, e.g., PCR. Typically, this involves the use of primersequences that are complementary to the two strands of the targetsequence both upstream and downstream from the polymorphism. AsymmetricPCR techniques may also be used. Amplified target, generallyincorporating a label, is then hybridized with the array underappropriate conditions. Upon completion of hybridization and washing ofthe array, the array is scanned to determine the position on the arrayto which the target sequence hybridizes. The hybridization data obtainedfrom the scan is typically in the form of fluorescence intensities as afunction of location on the array.

[0101] Although primarily described in terms of a single detectionblock, e.g., for detection of a single polymorphism, arrays can includemultiple detection blocks, and thus be capable of analyzing multiple,specific polymorphisms. In alternate arrangements, it will generally beunderstood that detection blocks may be grouped within a single array orin multiple, separate arrays so that varying, optimal conditions may beused during the hybridization of the target to the array. For example,it may often be desirable to provide for the detection of thosepolymorphisms that fall within G-C rich stretches of a genomic sequence,separately from those falling in A-T rich segments. This allows for theseparate optimization of hybridization conditions for each situation.

[0102] Additional description of use of oligonucleotide arrays fordetection of polymorphisms can be found, for example, in U.S. Pat. Nos.5,858,659 and 5,837,832, the entire teachings of which are incorporatedby reference herein.

[0103] Other methods of nucleic acid analysis can be used to detectpolymorphisms in MATN3 or splicing variants encoding by MATN3.Representative methods include direct manual sequencing (Church andGilbert (1988) Proc. Natl. Acad. Sci. USA 81:1991-1995; Sanger et al.(1977) Proc. Natl. Acad. Sci. 74:5463-5467; Beavis et al. U.S. Pat. No.5,288,644); automated fluorescent sequencing; single-strandedconformation polymorphism assays (SSCP); clamped denaturing gelelectrophoresis (CDGE); denaturing gradient gel electrophoresis (DGGE)(Sheffield et al. (1989) Proc. Natl. Acad. Sci. USA 86:232-236),mobility shift analysis (Orita et al. (1989) Proc. Natl. Acad. Sci. USA86:2766-2770), restriction enzyme analysis (Flavell et al. (1978) Cell15:25; Geever et al. (1981) Proc. Natl. Acad. Sci. USA 78:5081);heteroduplex analysis; chemical mismatch cleavage (CMC) (Cotton et al.(1985) Proc. Natl. Acad. Sci. USA 85:4397-4401); RNase protection assays(Myers et al. (1985) Science 230:1242); use of polypeptides whichrecognize nucleotide mismatches, such as E. coli mutS protein; andallele-specific PCR.

[0104] In one embodiment of the invention, diagnosis of a disease orcondition associated with a MATN3 nucleic acid (e.g., osteoarthritis) ora susceptibility to a disease or condition associated with a MATN3nucleic acid (e.g., osteoarthritis) can also be made by expressionanalysis by quantitative PCR (kinetic thermal cycling). This technique,utilizing TaqMan®, can be used to allow the identification ofpolymorphisms and whether a patient is homozygous or heterozygous. Thetechnique can assess the presence of an alteration in the expression orcomposition of the polypeptide encoded by a MATN3 nucleic acid orsplicing variants encoded by a MATN3 nucleic acid. Further, theexpression of the variants can be quantified as physically orfunctionally different.

[0105] In another embodiment of the invention, diagnosis of asusceptibility to osteoarthritis can also be made by examiningexpression and/or composition of a MATN3 polypeptide, by a variety ofmethods, including enzyme linked immunosorbent assays (ELISAs), Westernblots, immunoprecipitations and immunofluorescence. A test sample froman individual is assessed for the presence of an alteration in theexpression and/or an alteration in composition of the polypeptideencoded by MATN3, or for the presence of a particular variant encoded byMATN3. An alteration in expression of a polypeptide encoded by MATN3 canbe, for example, an alteration in the quantitative polypeptideexpression (i.e., the amount of polypeptide produced); an alteration inthe composition of a polypeptide encoded by MATN3 is an alteration inthe qualitative polypeptide expression (e.g., expression of a mutantMATN3 polypeptide or of a different splicing variant). In a preferredembodiment, diagnosis of a susceptibility to osteoarthritis is made bydetecting a particular splicing variant encoded by MATN3, or aparticular pattern of splicing variants.

[0106] Both such alterations (quantitative and qualitative) can also bepresent. An “alteration” in the polypeptide expression or composition,as used herein, refers to an alteration in expression or composition ina test sample, as compared with the expression or composition ofpolypeptide by MATN3 in a control sample. A control sample is a samplethat corresponds to the test sample (e.g., is from the same type ofcells), and is from an individual who is not affected by osteoarthritis.An alteration in the expression or composition of the polypeptide in thetest sample, as compared with the control sample, is indicative of asusceptibility to osteoarthritis. Similarly, the presence of one or moredifferent splicing variants in the test sample, or the presence ofsignificantly different amounts of different splicing variants in thetest sample, as compared with the control sample, is indicative of asusceptibility to osteoarthritis. Various means of examining expressionor composition of the polypeptide encoded by MATN3 can be used,including spectroscopy, colorimetry, electrophoresis, isoelectricfocusing, and immunoassays (e.g., David et al., U.S. Pat. No. 4,376,110)such as immunoblotting (see also Current Protocols in Molecular Biology,particularly chapter 10). For example, in one embodiment, an antibodycapable of binding to the polypeptide (e.g., as described above),preferably an antibody with a detectable label, can be used. Antibodiescan be polyclonal, or more preferably, monoclonal. An intact antibody,or a fragment thereof (e.g., Fab or F(ab′)₂) can be used. The term“labeled”, with regard to the probe or antibody, is intended toencompass direct labeling of the probe or antibody by coupling (i.e.,physically linking) a detectable substance to the probe or antibody, aswell as indirect labeling of the probe or antibody by reactivity withanother reagent that is directly labeled. Examples of indirect labelinginclude detection of a primary antibody using a fluorescently labeledsecondary antibody and end-labeling of a DNA probe with biotin such thatit can be detected with fluorescently labeled streptavidin.

[0107] Western blotting analysis, using an antibody as described abovethat specifically binds to a polypeptide encoded by a mutant MATN3, oran antibody that specifically binds to a polypeptide encoded by anon-mutant gene, or an antibody that specifically binds to a particularsplicing variant encoded by MATN3, can be used to identify the presencein a test sample of a particular splicing variant or of a polypeptideencoded by a polymorphic or mutant MATN3, or the absence in a testsample of a particular splicing variant or of a polypeptide encoded by anon-polymorphic or non-mutant gene. The presence of a polypeptideencoded by a polymorphic or mutant gene, or the absence of a polypeptideencoded by a non-polymorphic or non-mutant gene, is diagnostic for asusceptibility to osteoarthritis, as is the presence (or absence) ofparticular splicing variants encoded by the MATN3 nucleic acid.

[0108] In one embodiment of this method, the level or amount ofpolypeptide encoded by MATN3 in a test sample is compared with the levelor amount of the polypeptide encoded by MATN3 in a control sample. Alevel or amount of the polypeptide in the test sample that is higher orlower than the level or amount of the polypeptide in the control sample,such that the difference is statistically significant, is indicative ofan alteration in the expression of the polypeptide encoded by MATN3, andis diagnostic for a susceptibility to osteoarthritis. Alternatively, thecomposition of the polypeptide encoded by MATN3 in a test sample iscompared with the composition of the polypeptide encoded by MATN3 in acontrol sample (e.g., the presence of different splicing variants). Adifference in the composition of the polypeptide in the test sample, ascompared with the composition of the polypeptide in the control sample,is diagnostic for a susceptibility to osteoarthritis. In anotherembodiment, both the level or amount and the composition of thepolypeptide can be assessed in the test sample and in the controlsample. A difference in the amount or level of the polypeptide in thetest sample, compared to the control sample; a difference in compositionin the test sample, compared to the control sample; or both a differencein the amount or level, and a difference in the composition, isindicative of a susceptibility to osteoarthritis.

[0109] The invention also pertains to methods of diagnosing asusceptibility to osteoarthritis in an individual, comprising screeningfor an at-risk haplotype in the MATN3 gene that is more frequentlypresent in an individual susceptible to osteoarthritis (affected),compared to the frequency of its presence in a healthy individual(control), wherein the presence of the haplotype is indicative ofsusceptibility to osteoarthritis. Standard techniques for genotyping forthe presence of SNPs and/or microsatellite markers that are associatedwith osteoarthritis can be used, such as fluorescent based techniques(Chen et al. (1999) Genome Res. 9:492), PCR, LCR, Nested PCR, kineticthermal cycling to determine whether the patient is heterozygous orhomozygous for a particular genotype and other techniques for nucleicacid amplification. In a preferred embodiment, the method comprisesassessing in an individual the presence or frequency of SNPs and/ormicrosatellites in the MATN3 gene that are associated withosteoarthritis, wherein an excess or higher frequency of the SNPs and/ormicrosatellites compared to a healthy control individual is indicativethat the individual is susceptible to osteoarthritis.

[0110] See Table 3, FIGS. 5A-5C, Table 6 and Table 7 for SNPs andmarkers that comprise haplotypes that can be used as screening tools.SNPs and markers from these lists represent at-risk haplotypes and canbe used to design diagnostic tests for determining a susceptibility toosteoarthritis.

[0111] In one embodiment, the at-risk haplotype is characterized by thepresence of the polymorphism(s) represented by one or a combination ofsingle nucleotide polymorphisms at nucleic acid positions 58162, 57927,56822, 47929, 45434, 45317, 45178 and 45010, relative to SEQ ID NO: 1.In another embodiment, a diagnostic method for susceptibility toosteoarthritis can comprise determining the presence of at-riskhaplotype represented by one or a combination of single nucleotidepolymorphisms and microsatellie markers at nucleic acid positions 58162,57927, 56822, 47929, 45434, 45317, 45178 and 45010, relative to SEQ IDNO: 1.

[0112] Kits (e.g., reagent kits) useful in the methods of diagnosiscomprise components useful in any of the methods described herein,including, for example, hybridization probes or primers as describedherein (e.g., labeled probes or primers), reagents for detection oflabeled molecules, restriction enzymes (e.g., for RFLP analysis),allele-specific oligonucleotides, antibodies which bind to altered or tonon-altered (native) MATN3 polypeptide, means for amplification ofnucleic acids comprising MATN3, or means for analyzing the nucleic acidsequence of MATN3 or for analyzing the amino acid sequence of a MATN3polypeptide, etc. In one embodiment, a kit for diagnosing susceptibilityto osteoarthritis can comprise primers for nucleic acid amplification ofa region in the MATN3 gene comprising an at-risk haplotype that is morefrequently present in an individual susceptible to osteoarthritis. Theprimers can be designed using portions of the nucleic acids flankingSNPs that are indicative of osteoarthritis. In a particularly preferredembodiment, the primers are designed to amplify regions of the MATN3gene associated with an at-risk haplotype for osteoarthritis, shown inTables 6 and 7. In another embodiment of the invention, a kit fordiagnosing susceptibility to osteoarthritis can further comprise probesdesigned to hybridize to regions of the MATN3 gene associated with anat-risk haplotype for osteoarthritis, shown in, for example, Table 6 andTable 7. The at risk haplotype can be characterized, for example, by thepresence of at least one single nucleotide polymorphism at nucleic acidpositions 58162, 57927, 56822, 47929, 45434, 45317, 45178 and 45010,relative to SEQ ID NO: 1.

[0113] In another embodiment of the invention, a method for thediagnosis and identification of susceptibility to osteoarthritis in anindividual is provided by identifying an at-risk haplotype in MATN3. Inone embodiment, the at-risk haplotype is a haplotype for which thepresence of the haplotype increases the risk of osteoarthritissignificantly. Although it is to be understood that identifying whethera risk is significant may depend on a variety of factors, including thespecific disease, the haplotype, and often, environmental factors, thesignificance may be measured by an odds ratio or a percentage. In oneembodiment, a significant risk is measured as an odds ratio of at leastabout 1.1, including but not limited to: 1.2, 1.3, 1.4, 1.5, 1.6, 1.7,1.8 and 1.9. In a further embodiment, an odds ratio of at least 1.2 issignificant. In a further embodiment, an odds ratio of at least about1.5 is significant. In another embodiment, an odds ratio of at leastabout 1.7 is significant. In a further embodiment, a significantincrease in risk is at least about 20%, including but not limited to:25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%,95% and 98%.

[0114] Screening Assays and Agents Identified Thereby

[0115] The invention provides methods (also referred to herein as“screening assays”) for identifying the presence of a nucleotide thathybridizes to a nucleic acid of the invention, as well as foridentifying the presence of a polypeptide encoded by a nucleic acid ofthe invention. In one embodiment, the presence (or absence) of a nucleicacid molecule of interest (e.g., a nucleic acid that has significanthomology with a nucleic acid of the invention) in a sample can beassessed by contacting the sample with a nucleic acid comprising anucleic acid of the invention (e.g., a nucleic acid having the sequenceof SEQ ID NO: 1 and comprising at least one polymorphism shown in Table3 or FIGS. 5A-5C, or the complements thereof, or a nucleic acid encodingan amino acid having the sequence of SEQ ID NO: 2, 3, 4, 5, 6, 7, 8 or9, or a fragment or variant of such nucleic acids), under stringentconditions as described above, and then assessing the sample for thepresence (or absence) of hybridization. In a preferred embodiment, highstringency conditions are conditions appropriate for selectivehybridization. In another embodiment, a sample containing the nucleicacid molecule of interest is contacted with a nucleic acid containing acontiguous nucleotide sequence (e.g., a primer or a probe as describedabove) that is at least partially complementary to a part of the nucleicacid molecule of interest (e.g., a MATN3 nucleic acid), and thecontacted sample is assessed for the presence or absence ofhybridization. In a preferred embodiment, the nucleic acid containing acontiguous nucleotide sequence is completely complementary to a part ofthe nucleic acid molecule of interest.

[0116] In any of these embodiments, all or a portion of the nucleic acidof interest can be subjected to amplification prior to performing thehybridization.

[0117] In another embodiment, the presence (or absence) of a polypeptideof interest, such as a polypeptide of the invention or a fragment orvariant thereof, in a sample can be assessed by contacting the samplewith an antibody that specifically binds to the polypeptide of interest(e.g., an antibody such as those described above), and then assessingthe sample for the presence (or absence) of binding of the antibody tothe polypeptide of interest.

[0118] In another embodiment, the invention provides methods foridentifying agents (e.g., fusion proteins, polypeptides,peptidomimetics, prodrugs, receptors, binding agents, antibodies, smallmolecules or other drugs, or ribozymes which alter (e.g., increase ordecrease) the activity of the polypeptides described herein, or whichotherwise interact with the polypeptides herein. For example, suchagents can be agents which bind to polypeptides described herein (e.g.,MATN3 binding agents); which have a stimulatory or inhibitory effect on,for example, activity of polypeptides of the invention; or which change(e.g., enhance or inhibit) the ability of the polypeptides of theinvention to interact with MATN3 binding agents (e.g., receptors orother binding agents); or which alter posttranslational processing ofthe MATN3 polypeptide (e.g., agents that alter proteolytic processing todirect the polypeptide from where it is normally synthesized to anotherlocation in the cell, such as the cell surface; agents that alterproteolytic processing such that more polypeptide is released from thecell, etc.).

[0119] In one embodiment, the invention provides assays for screeningcandidate or test agents that bind to or modulate the activity ofpolypeptides described herein (or biologically active portion(s)thereof), as well as agents identifiable by the assays. Test agents canbe obtained using any of the numerous approaches in combinatoriallibrary methods known in the art, including: biological libraries;spatially addressable parallel solid phase or solution phase libraries;synthetic library methods requiring deconvolution; the “one-beadone-compound” library method; and synthetic library methods usingaffinity chromatography selection. The biological library approach islimited to polypeptide libraries, while the other four approaches areapplicable to polypeptide, non-peptide oligomer or small moleculelibraries of compounds (Lam, K. S. (1997) Anticancer Drug Des. 12:145).

[0120] In one embodiment, to identify agents which alter the activity ofa MATN3 polypeptide, a cell, cell lysate, or solution containing orexpressing a MATN3 polypeptide (e.g., SEQ ID NO: 2, 3, 4, 5, 6, 7, 8 or9, or another splicing variant encoded by MATN3), or a fragment orderivative thereof (as described above), can be contacted with an agentto be tested; alternatively, the polypeptide can be contacted directlywith the agent to be tested. The level (amount) of MATN3 activity isassessed (e.g., the level (amount) of MATN3 activity is measured, eitherdirectly or indirectly), and is compared with the level of activity in acontrol (i.e., the level of activity of the MATN3 polypeptide or activefragment or derivative thereof in the absence of the agent to betested). If the level of the activity in the presence of the agentdiffers, by an amount that is statistically significant, from the levelof the activity in the absence of the agent, then the agent is an agentthat alters the activity of MATN3 polypeptide. An increase in the levelof MATN3 activity relative to a control, indicates that the agent is anagent that enhances (is an agonist of) MATN3 activity. Similarly, adecrease in the level of MATN3 activity relative to a control, indicatesthat the agent is an agent that inhibits (is an antagonist of) MATN3activity. In another embodiment, the level of activity of a MATN3polypeptide or derivative or fragment thereof in the presence of theagent to be tested, is compared with a control level that has previouslybeen established. A level of the activity in the presence of the agentthat differs from the control level by an amount that is statisticallysignificant indicates that the agent alters MATN3 activity.

[0121] The present invention also relates to an assay for identifyingagents which alter the expression of the MATN3 nucleic acid (e.g.,antisense nucleic acids, fusion proteins, polypeptides, peptidomimetics,prodrugs, receptors, binding agents, antibodies, small molecules orother drugs, or ribozymes) which alter (e.g., increase or decrease)expression (e.g., transcription or translation) of the gene or whichotherwise interact with the nucleic acids described herein, as well asagents identifiable by the assays. For example, a solution containing anucleic acid encoding MATN3 polypeptide (e.g., MATN3 nucleic acid) canbe contacted with an agent to be tested. The solution can comprise, forexample, cells containing the nucleic acid or cell lysate containing thenucleic acid; alternatively, the solution can be another solution whichcomprises elements necessary for transcription/translation of thenucleic acid. Cells not suspended in solution can also be employed, ifdesired. The level and/or pattern of MATN3 expression (e.g., the leveland/or pattern of mRNA or of protein expressed, such as the level and/orpattern of different splicing variants) is assessed, and is comparedwith the level and/or pattern of expression in a control (i.e., thelevel and/or pattern of the MATN3 expression in the absence of the agentto be tested). If the level and/or pattern in the presence of the agentdiffers, by an amount or in a manner that is statistically significant,from the level and/or pattern in the absence of the agent, then theagent is an agent that alters the expression of MATN3. Enhancement ofMATN3 expression indicates that the agent is an agonist of MATN3activity. Similarly, inhibition of MATN3 expression indicates that theagent is an antagonist of MATN3 activity. In another embodiment, thelevel and/or pattern of MATN3 polypeptide(s) (e.g., different splicingvariants) in the presence of the agent to be tested, is compared with acontrol level and/or pattern that has previously been established. Alevel and/or pattern in the presence of the agent that differs from thecontrol level and/or pattern by an amount or in a manner that isstatistically significant indicates that the agent alters MATN3expression.

[0122] In another embodiment of the invention, agents which alter theexpression of the MATN3 nucleic acid or which otherwise interact withthe nucleic acids described herein, can be identified using a cell, celllysate, or solution containing a nucleic acid encoding the promoterregion of the MATN3 gene operably linked to a reporter gene. Aftercontact with an agent to be tested, the level of expression of thereporter gene (e.g., the level of mRNA or of protein expressed) isassessed, and is compared with the level of expression in a control(i.e., the level of the expression of the reporter gene in the absenceof the agent to be tested). If the level in the presence of the agentdiffers, by an amount or in a manner that is statistically significant,from the level in the absence of the agent, then the agent is an agentthat alters the expression of MATN3, as indicated by its ability toalter expression of a gene that is operably linked to the MATN3 genepromoter. Enhancement of the expression of the reporter indicates thatthe agent is an agonist of MATN3 activity. Similarly, inhibition of theexpression of the reporter indicates that the agent is an antagonist ofMATN3 activity. In another embodiment, the level of expression of thereporter in the presence of the agent to be tested, is compared with acontrol level that has previously been established. A level in thepresence of the agent that differs from the control level by an amountor in a manner that is statistically significant indicates that theagent alters MATN3 expression.

[0123] Agents which alter the amounts of different splicing variantsencoded by MATN3 (e.g., an agent which enhances activity of a firstsplicing variant, and which inhibits activity of a second splicingvariant), as well as agents which are agonists of activity of a firstsplicing variant and antagonists of activity of a second splicingvariant, can easily be identified using these methods described above.

[0124] In other embodiments of the invention, assays can be used toassess the impact of a test agent on the activity of a polypeptide inrelation to a MATN3 binding agent. For example, a cell that expresses acompound that interacts with MATN3 (herein referred to as a “MATN3binding agent”, which can be a polypeptide or other molecule thatinteracts with MATN3, such as a receptor) is contacted with MATN3 in thepresence of a test agent, and the ability of the test agent to alter theinteraction between MATN3 and the MATN3 binding agent is determined.Alternatively, a cell lysate or a solution containing the MATN3 bindingagent, can be used. An agent which binds to MATN3 or the MATN3 bindingagent can alter the interaction by interfering with, or enhancing theability of MATN3 to bind to, associate with, or otherwise interact withthe MATN3 binding agent. Determining the ability of the test agent tobind to MATN3 or a MATN3 binding agent can be accomplished, for example,by coupling the test agent with a radioisotope or enzymatic label suchthat binding of the test agent to the polypeptide can be determined bydetecting the labeled with ¹²⁵I, ³⁵S, ¹⁴C or ³H, either directly orindirectly, and the radioisotope detected by direct counting ofradioemmission or by scintillation counting. Alternatively, test agentscan be enzymatically labeled with, for example, horseradish peroxidase,alkaline phosphatase, or luciferase, and the enzymatic label detected bydetermination of conversion of an appropriate substrate to product. Itis also within the scope of this invention to determine the ability of atest agent to interact with the polypeptide without the labeling of anyof the interactants. For example, a microphysiometer can be used todetect the interaction of a test agent with MATN3 or a MATN3 bindingagent without the labeling of either the test agent, MATN3, or the MATN3binding agent. McConnell, H. M. et al. (1992) Science 257:1906-1912. Asused herein, a “microphysiometer” (e.g., Cytosensor™) is an analyticalinstrument that measures the rate at which a cell acidifies itsenvironment using a light-addressable potentiometric sensor (LAPS).Changes in this acidification rate can be used as an indicator of theinteraction between ligand and polypeptide. See the ExemplificationSection for a discussion of known MATN3 binding partners. Thus, thesereceptors can be used to screen for compounds that are MATN3 receptoragonists for use in treating osteoarthritis or MATN3 receptorantagonists for studying osteoarthritis. The linkage data providedherein, for the first time, provides such correction to osteoarthritis.Drugs could be designed to regulate MATN3 receptor activation which inturn can be used to regulate signaling pathways and transcription eventsof genes downstream, such as Cbfa1.

[0125] In another embodiment of the invention, assays can be used toidentify polypeptides that interact with one or more MATN3 polypeptides,as described herein. For example, a yeast two-hybrid system such as thatdescribed by Fields and Song (Fields and Song (1989) Nature 340:245-246)can be used to identify polypeptides that interact with one or moreMATN3 polypeptides. In such a yeast two-hybrid system, vectors areconstructed based on the flexibility of a transcription factor which hastwo functional domains (a DNA binding domain and a transcriptionactivation domain). If the two domains are separated but fused to twodifferent proteins that interact with one another, transcriptionalactivation can be achieved, and transcription of specific markers (e.g.,nutritional markers such as His and Ade, or color markers such as lacZ)can be used to identify the presence of interaction and transcriptionalactivation. For example, in the methods of the invention, a first vectoris used which includes a nucleic acid encoding a DNA binding domain andalso a MATN3 polypeptide, splicing variant, or fragment or derivativethereof, and a second vector is used which includes a nucleic acidencoding a transcription activation domain and also a nucleic acidencoding a polypeptide which potentially may interact with the MATN3polypeptide, splicing variant, or fragment or derivative thereof (e.g.,a MATN3 polypeptide binding agent or receptor). Incubation of yeastcontaining the first vector and the second vector under appropriateconditions (e.g., mating conditions such as used in the Matchmaker™system from Clontech) allows identification of colonies which expressthe markers of interest. These colonies can be examined to identify thepolypeptide(s) which interact with the MATN3 polypeptide or fragment orderivative thereof. Such polypeptides may be useful as agents whichalter the activity of expression of a MATN3 polypeptide, as describedabove.

[0126] In more than one embodiment of the above assay methods of thepresent invention, it may be desirable to immobilize either MATN3, theMATN3 binding agent, or other components of the assay on a solidsupport, in order to facilitate separation of complexed from uncomplexedforms of one or both of the polypeptides, as well as to accommodateautomation of the assay. Binding of a test agent to the polypeptide, orinteraction of the polypeptide with a binding agent in the presence andabsence of a test agent, can be accomplished in any vessel suitable forcontaining the reactants. Examples of such vessels include microtitreplates, test tubes, and micro-centrifuge tubes. In one embodiment, afusion protein (e.g., a glutathione-S-transferase fusion protein) can beprovided which adds a domain that allows MATN3 or a MATN3 binding agentto be bound to a matrix or other solid support.

[0127] In another embodiment, modulators of expression of nucleic acidmolecules of the invention are identified in a method wherein a cell,cell lysate, or solution containing a nucleic acid encoding MATN3 iscontacted with a test agent and the expression of appropriate mRNA orpolypeptide (e.g., splicing variant(s)) in the cell, cell lysate, orsolution, is determined. The level of expression of appropriate mRNA orpolypeptide(s) in the presence of the test agent is compared to thelevel of expression of mRNA or polypeptide(s) in the absence of the testagent. The test agent can then be identified as a modulator ofexpression based on this comparison. For example, when expression ofmRNA or polypeptide is greater (statistically significantly greater) inthe presence of the test agent than in its absence, the test agent isidentified as a stimulator or enhancer of the mRNA or polypeptideexpression. Alternatively, when expression of the mRNA or polypeptide isless (statistically significantly less) in the presence of the testagent than in its absence, the test agent is identified as an inhibitorof the mRNA or polypeptide expression. The level of mRNA or polypeptideexpression in the cells can be determined by methods described hereinfor detecting mRNA or polypeptide.

[0128] This invention further pertains to novel agents identified by theabove-described screening assays. Accordingly, it is within the scope ofthis invention to further use an agent identified as described herein inan appropriate animal model. For example, an agent identified asdescribed herein (e.g., a test agent that is a modulating agent, anantisense nucleic acid molecule, a specific antibody, or apolypeptide-binding agent) can be used in an animal model to determinethe efficacy, toxicity, or side effects of treatment with such an agent.Alternatively, an agent identified as described herein can be used in ananimal model to determine the mechanism of action of such an agent.Furthermore, this invention pertains to uses of novel agents identifiedby the above-described screening assays for treatments as describedherein. In addition, an agent identified as described herein can be usedto alter activity of a polypeptide encoded by MATN3, or to alterexpression of MATN3, by contacting the polypeptide or the gene (orcontacting a cell comprising the polypeptide or the gene) with the agentidentified as described herein.

[0129] Pharmaceutical Compositions

[0130] The present invention also pertains to pharmaceuticalcompositions comprising nucleic acids described herein, particularlynucleotides encoding the polypeptides described herein; comprising thenormal (not associated with osteoarthritis) MATN 3 gene product,polypeptides described herein (e.g., one or more of SEQ ID NO: 2, 3, 4,5, 6, 7, 8 or 9); and/or comprising other splicing variants encoded byMATN3; and/or an agent that alters (e.g., enhances or inhibits) MATN3gene expression or MATN3 polypeptide activity as described herein. Forinstance, a polypeptide, protein (e.g., a MATN3 receptor), an agent thatalters MATN3 gene expression, or a MATN3 binding agent or bindingpartner, fragment, fusion protein or prodrug thereof, or a nucleotide ornucleic acid construct (vector) comprising a nucleotide of the presentinvention, or an agent that alters MATN3 polypeptide activity, can beformulated with a physiologically acceptable carrier or excipient toprepare a pharmaceutical composition. The carrier and composition can besterile. The formulation should suit the mode of administration.

[0131] Suitable pharmaceutically acceptable carriers include but are notlimited to water, salt solutions (e.g., NaCl), saline, buffered saline,alcohols, glycerol, ethanol, gum arabic, vegetable oils, benzylalcohols, polyethylene glycols, gelatin, carbohydrates such as lactose,amylose or starch, dextrose, magnesium stearate, talc, silicic acid,viscous paraffin, perfume oil, fatty acid esters,hydroxymethylcellulose, polyvinyl pyro-lidone, etc., as well ascombinations thereof. The pharmaceutical preparations can, if desired,be mixed with auxiliary agents, e.g., lubricants, preservatives,stabilizers, wetting agents, emulsifiers, salts for influencing osmoticpressure, buffers, coloring, flavoring and/or aromatic substances andthe like which do not deleteriously react with the active agents.

[0132] The composition, if desired, can also contain minor amounts ofwetting or emulsifying agents, or pH buffering agents. The compositioncan be a liquid solution, suspension, emulsion, tablet, pill, capsule,sustained release formulation, or powder. The composition can beformulated as a suppository, with traditional binders and carriers suchas triglycerides. Oral formulation can include standard carriers such aspharmaceutical grades of mannitol, lactose, starch, magnesium stearate,polyvinyl pyrollidone, sodium saccharine, cellulose, magnesiumcarbonate, etc.

[0133] Methods of introduction of these compositions include, but arenot limited to, intradermal, intramuscular, intraperitoneal,intraocular, intravenous, subcutaneous, topical, oral and intranasal.Other suitable methods of introduction can also include gene therapy (asdescribed below), rechargeable or biodegradable devices, particleacceleration devices (“gene guns”) and slow release polymeric devices.The pharmaceutical compositions of this invention can also beadministered as part of a combinatorial therapy with other agents.

[0134] The composition can be formulated in accordance with the routineprocedures as a pharmaceutical composition adapted for administration tohuman beings. For example, compositions for intravenous administrationtypically are solutions in sterile isotonic aqueous buffer. Wherenecessary, the composition may also include a solubilizing agent and alocal anesthetic to ease pain at the site of the injection. Generally,the ingredients are supplied either separately or mixed together in unitdosage form, for example, as a dry lyophilized powder or water freeconcentrate in a hermetically sealed container such as an ampule orsachette indicating the quantity of active agent. Where the compositionis to be administered by infusion, it can be dispensed with an infusionbottle containing sterile pharmaceutical grade water, saline ordextrose/water. Where the composition is administered by injection, anampule of sterile water for injection or saline can be provided so thatthe ingredients may be mixed prior to administration.

[0135] For topical application, non-sprayable forms, viscous tosemi-solid or solid forms comprising a carrier compatible with topicalapplication and having a dynamic viscosity preferably greater thanwater, can be employed. Suitable formulations include but are notlimited to solutions, suspensions, emulsions, creams, ointments,powders, enemas, lotions, sols, liniments, salves, aerosols, etc., whichare, if desired, sterilized or mixed with auxiliary agents, e.g.,preservatives, stabilizers, wetting agents, buffers or salts forinfluencing osmotic pressure, etc. The agent may be incorporated into acosmetic formulation. For topical application, also suitable aresprayable aerosol preparations wherein the active ingredient, preferablyin combination with a solid or liquid inert carrier material, ispackaged in a squeeze bottle or in admixture with a pressurizedvolatile, normally gaseous propellant, e.g., pressurized air.

[0136] Agents described herein can be formulated as neutral or saltforms. Pharmaceutically acceptable salts include those formed with freeamino groups such as those derived from hydrochloric, phosphoric,acetic, oxalic, tartaric acids, etc., and those formed with freecarboxyl groups such as those derived from sodium, potassium, ammonium,calcium, ferric hydroxides, isopropylamine, triethylamine, 2-ethylaminoethanol, histidine, procaine, etc.

[0137] The agents are administered in a therapeutically effectiveamount. The amount of agents which will be therapeutically effective inthe treatment of a particular disorder or condition will depend on thenature of the disorder or condition, and can be determined by standardclinical techniques. In addition, in vitro or in vivo assays mayoptionally be employed to help identify optimal dosage ranges. Theprecise dose to be employed in the formulation will also depend on theroute of administration, and the seriousness of the symptoms ofosteoarthritis, and should be decided according to the judgment of apractitioner and each patient's circumstances. Effective doses may beextrapolated from dose-response curves derived from in vitro or animalmodel test systems.

[0138] The invention also provides a pharmaceutical pack or kitcomprising one or more containers filled with one or more of theingredients of the pharmaceutical compositions of the invention.Optionally associated with such container(s) can be a notice in the formprescribed by a governmental agency regulating the manufacture, use orsale of pharmaceuticals or biological products, which notice reflectsapproval by the agency of manufacture, use of sale for humanadministration. The pack or kit can be labeled with informationregarding mode of administration, sequence of drug administration (e.g.,separately, sequentially or concurrently), or the like. The pack or kitmay also include means for reminding the patient to take the therapy.The pack or kit can be a single unit dosage of the combination therapyor it can be a plurality of unit dosages. In particular, the agents canbe separated, mixed together in any combination, present in a singlevial or tablet. Agents assembled in a blister pack or other dispensingmeans is preferred. For the purpose of this invention, unit dosage isintended to mean a dosage that is dependent on the individualpharmacodynamics of each agent and administered in FDA approved dosagesin standard time courses.

[0139] Methods of Therapy

[0140] The present invention also pertains to methods of treatment(prophylactic and/or therapeutic) for osteoarthritis using a MATN3therapeutic agent. A “MATN3 therapeutic agent” is an agent that alters(e.g., enhances or inhibits) MATN3 polypeptide activity and/or MATN3nucleic acid expression, as described herein (e.g., a MATN3 agonist orantagonist). MATN3 therapeutic agents can alter MATN3 polypeptideactivity or gene expression by a variety of means, such as, for example,by providing additional MATN3 polypeptide or by upregulating thetranscription or translation of the MATN3 nucleic acid; by alteringposttranslational processing of the MATN3 polypeptide; by alteringtranscription of MATN3 splicing variants; or by interfering with MATN3polypeptide activity (e.g., by binding to a MATN3 polypeptide), or bydownregulating the transcription or translation of the MATN3 nucleicacid. Representative MATN3 therapeutic agents include the following:nucleic acids or fragments or derivatives thereof described herein,particularly nucleotides encoding the polypeptides described herein andvectors comprising such nucleic acids (e.g., a gene, cDNA, and/or mRNA,such as a nucleic acid encoding a MATN3 polypeptide or active fragmentor derivative thereof, or an oligonucleotide; for example, non-alteredMATN3); polypeptides described herein (e.g., non-altered MATN3); otherpolypeptides (e.g., MATN3 receptors); MATN3 binding agents;peptidomimetics; fusion proteins or prodrugs thereof, antibodies (e.g.,an antibody to an altered MATN3 polypeptide, or an antibody to anon-altered MATN3 polypeptide, or an antibody to a particular splicingvariant encoded by MATN3, as described above); ribozymes; other smallmolecules; and other agents that alter (e.g., enhance or inhibit) MATN3gene expression or polypeptide activity, or that regulate transcriptionof MATN3 splicing variants (e.g., agents that affect which splicingvariants are expressed, or that affect the amount of each splicingvariant that is expressed.

[0141] More than one MATN3 therapeutic agent can be used concurrently,if desired.

[0142] The MATN3 therapeutic agent that is a nucleic acid is used in thetreatment of osteoarthritis. The term, “treatment” as used herein,refers not only to ameliorating symptoms associated with the disease,but also preventing or delaying the onset of the disease, and alsolessening the severity or frequency of symptoms of the disease. Thetherapy is designed to alter (e.g., inhibit or enhance), replace orsupplement activity of a MATN3 polypeptide in an individual. Forexample, a MATN3 therapeutic agent can be administered in order toupregulate or increase the expression or availability of the MATN3 geneor of specific splicing variants of MATN3, or, conversely, todownregulate or decrease the expression or availability of the MATN3gene or specific splicing variants of MATN3. Upregulation or increasingexpression or availability of a native MATN3 gene or of a particularsplicing variant could interfere with or compensate for the expressionor activity of a defective gene or another splicing variant;downregulation or decreasing expression or availability of a nativeMATN3 gene or of a particular splicing variant could minimize theexpression or activity of a defective gene or the particular splicingvariant and thereby minimize the impact of the defective gene or theparticular splicing variant.

[0143] The MATN3 therapeutic agent(s) are administered in atherapeutically effective amount (i.e., an amount that is sufficient totreat the disease, such as by ameliorating symptoms associated with thedisease, preventing or delaying the onset of the disease, and/or alsolessening the severity or frequency of symptoms of the disease). Theamount which will be therapeutically effective in the treatment of aparticular individual's disorder or condition will depend on thesymptoms and severity of the disease, and can be determined by standardclinical techniques. In addition, in vitro or in vivo assays mayoptionally be employed to help identify optimal dosage ranges. Theprecise dose to be employed in the formulation will also depend on theroute of administration, and the seriousness of the disease or disorder,and should be decided according to the judgment of a practitioner andeach patient's circumstances. Effective doses may be extrapolated fromdose-response curves derived from in vitro or animal model test systems.

[0144] In one embodiment, a nucleic acid of the invention (e.g., anucleic acid encoding a MATN3 polypeptide, such as SEQ ID NO: 1 oranother nucleic acid that encodes a MATN3 polypeptide, derivative orfragment thereof) can be used, either alone or in a pharmaceuticalcomposition as described above. For example, MATN3 or a cDNA encodingthe MATN3 polypeptide, either by itself or included within a vector, canbe introduced into cells (either in vitro or in vivo) such that thecells produce native MATN3 polypeptide. If necessary, cells that havebeen transformed with the gene or cDNA or a vector comprising the geneor CDNA can be introduced (or re-introduced) into an individual affectedwith the disease. Thus, cells which, in nature, lack native MATN3expression and activity, or have mutant MATN3 expression and activity,or have expression of a disease-associated MATN3 splicing variant, canbe engineered to express MATN3 polypeptide or an active fragment of theMATN3 polypeptide (or a different variant of MATN3 polypeptide). In apreferred embodiment, nucleic acid encoding the MATN3 polypeptide, or anactive fragment or derivative thereof, can be introduced into anexpression vector, such as a viral vector, and the vector can beintroduced into appropriate cells in an animal. Other gene transfersystems, including viral and nonviral transfer systems, can be used.Alternatively, nonviral gene transfer methods, such as calcium phosphatecoprecipitation, mechanical techniques (e.g., microinjection); membranefusion-mediated transfer via liposomes; or direct DNA uptake, can alsobe used.

[0145] Alternatively, in another embodiment of the invention, a nucleicacid of the invention; a nucleic acid complementary to a nucleic acid ofthe invention; or a portion of such a nucleic acid (e.g., anoligonucleotide as described below), can be used in “antisense” therapy,in which a nucleic acid (e.g., an oligonucleotide) which specificallyhybridizes to the mRNA and/or genomic DNA of MATN3 is administered orgenerated in situ. The antisense nucleic acid that specificallyhybridizes to the mRNA and/or DNA inhibits expression of the MATN3polypeptide, e.g., by inhibiting translation and/or transcription.Binding of the antisense nucleic acid can be by conventional base paircomplementarity, or, for example, in the case of binding to DNAduplexes, through specific interaction in the major groove of the doublehelix.

[0146] An antisense construct of the present invention can be delivered,for example, as an expression plasmid as described above. When theplasmid is transcribed in the cell, it produces RNA which iscomplementary to a portion of the mRNA and/or DNA which encodes MATN3polypeptide. Alternatively, the antisense construct can be anoligonucleotide probe which is generated ex vivo and introduced intocells; it then inhibits expression by hybridizing with the mRNA and/orgenomic DNA of MATN3. In one embodiment, the oligonucleotide probes aremodified oligonucleotides which are resistant to endogenous nucleases,e.g., exonucleases and/or endonucleases, thereby rendering them stablein vivo. Exemplary nucleic acid molecules for use as antisenseoligonucleotides are phosphoramidate, phosphothioate andmethylphosphonate analogs of DNA (see also U.S. Pat. Nos. 5,176,996;5,264,564; and 5,256,775). Additionally, general approaches toconstructing oligomers useful in antisense therapy are also described,for example, by Van der Krol et al. ((1988) Biotechniques 6:958-976);and Stein et al. ((1988) Cancer Res 48:2659-2668). With respect toantisense DNA, oligodeoxyribonucleotides derived from the translationinitiation site, e.g., between the −10 and +10 regions of MATN3sequence, are preferred.

[0147] To perform antisense therapy, oligonucleotides (mRNA, cDNA orDNA) are designed that are complementary to mRNA encoding MATN3. Theantisense oligonucleotides bind to MATN3 mRNA transcripts and preventtranslation. Absolute complementarity, although preferred, is notrequired. A sequence “complementary” to a portion of an RNA, as referredto herein, indicates that a sequence has sufficient complementarity tobe able to hybridize with the RNA, forming a stable duplex; in the caseof double-stranded antisense nucleic acids, a single strand of theduplex DNA may thus be tested, or triplex formation may be assayed. Theability to hybridize will depend on both the degree of complementarityand the length of the antisense nucleic acid, as described in detailabove. Generally, the longer the hybridizing nucleic acid, the more basemismatches with an RNA it may contain and still form a stable duplex (ortriplex, as the case may be). One skilled in the art can ascertain atolerable degree of mismatch by use of standard procedures.

[0148] The oligonucleotides used in antisense therapy can be DNA, RNA,or chimeric mixtures or derivatives or modified versions thereof,single-stranded or double-stranded. The oligonucleotides can be modifiedat the base moiety, sugar moiety, or phosphate backbone, for example, toimprove stability of the molecule, hybridization, etc. Theoligonucleotides can include other appended groups such as peptides(e.g., for targeting host cell receptors in vivo), or agentsfacilitating transport across the cell membrane (see, e.g., Letsinger etal. (1989) Proc. Natl. Acad. Sci. USA 86:6553-6556; Lemaitre et al.(1987) Proc. Natl. Acad Sci. USA 84:648-652; PCT InternationalPublication No. WO 88/09810) or the blood-brain barrier (see, e.g., PCTInternational Publication No. WO 89/10134), or hybridization-triggeredcleavage agents (see, e.g., Krol et al. (1988) BioTechniques 6:958-976)or intercalating agents. (See, e.g., Zon (1988), Pharm. Res. 5:539-549).To this end, the oligonucleotide may be conjugated to another molecule(e.g., a peptide, hybridization triggered cross-linking agent, transportagent, hybridization-triggered cleavage agent).

[0149] The antisense molecules are delivered to cells which expressMATN3 in vivo. A number of methods can be used for delivering antisenseDNA or RNA to cells; e.g., antisense molecules can be injected directlyinto the tissue site, or modified antisense molecules, designed totarget the desired cells (e.g., antisense linked to peptides orantibodies that specifically bind receptors or antigens expressed on thetarget cell surface) can be administered systemically. Alternatively, ina preferred embodiment, a recombinant DNA construct is utilized in whichthe antisense oligonucleotide is placed under the control of a strongpromoter (e.g., pol III or pol II). The use of such a construct totransfect target cells in the patient results in the transcription ofsufficient amounts of single stranded RNAs that will form complementarybase pairs with the endogenous MATN3 transcripts and thereby preventtranslation of the MATN3 mRNA. For example, a vector can be introducedin vivo such that it is taken up by a cell and directs the transcriptionof an antisense RNA. Such a vector can remain episomal or becomechromosomally integrated, as long as it can be transcribed to producethe desired antisense RNA. Such vectors can be constructed byrecombinant DNA technology methods standard in the art and describedabove. For example, a plasmid, cosmid, YAC or viral vector can be usedto prepare the recombinant DNA construct which can be introduceddirectly into the tissue site. Alternatively, viral vectors can be usedwhich selectively infect the desired tissue, in which caseadministration may be accomplished by another route (e.g.,systemically).

[0150] Endogenous MATN3 expression can also be reduced by inactivatingor “knocking out” MATN3 or its promoter using targeted homologousrecombination (e.g., see Smithies et al. (1985) Nature 317:230-234;Thomas and Capecchi (1987) Cell 51:503-512; Thompson et al. (1989) Cell5:313-321). For example, a mutant, non-functional MATN3 (or a completelyunrelated DNA sequence) flanked by DNA homologous to the endogenousMATN3 (either the coding regions or regulatory regions of MATN3) can beused, with or without a selectable marker and/or a negative selectablemarker, to transfect cells that express MATN3 in vivo. Insertion of theDNA construct, via targeted homologous recombination, results ininactivation of MATN3. The recombinant DNA constructs can be directlyadministered or targeted to the required site in vivo using appropriatevectors, as described above. Alternatively, expression of non-mutantMATN3 can be increased using a similar method: targeted homologousrecombination can be used to insert a DNA construct comprising anon-mutant, functional MATN3 (e.g., a gene having SEQ ID NO: 1 which mayoptionally comprise at least one polymorphism shown in Table 3 or FIGS.5A-5C), or a portion thereof, in place of a mutant MATN3 in the cell, asdescribed above. In another embodiment, targeted homologousrecombination can be used to insert a DNA construct comprising a nucleicacid that encodes a MATN3 polypeptide variant that differs from thatpresent in the cell.

[0151] Alternatively, endogenous MATN3 expression can be reduced bytargeting deoxyribonucleotide sequences complementary to the regulatoryregion of MATN3 (i.e., the MATN3 promoter and/or enhancers) to formtriple helical structures that prevent transcription of MATN3 in targetcells in the body. (See generally, Helene (1991) Anticancer Drug Des.6(6):569-84; Helene et al. (1992) Ann. N.Y. Acad. Sci. 660:27-36; andMaher, L. J. (1992) Bioassays 14(12):807-15). Likewise, the antisenseconstructs described herein, by antagonizing the normal biologicalactivity of one of the MATN3 proteins, can be used in the manipulationof tissue, e.g., tissue differentiation, both in vivo and for ex vivotissue cultures. Furthermore, the anti-sense techniques (e.g.,microinjection of antisense molecules, or transfection with plasmidswhose transcripts are anti-sense with regard to a MATN3 mRNA or genesequence) can be used to investigate role of MATN3 in developmentalevents, as well as the normal cellular function of MATN3 in adulttissue. Such techniques can be utilized in cell culture, but can also beused in the creation of transgenic animals.

[0152] In yet another embodiment of the invention, other MATN3therapeutic agents as described herein can also be used in the treatmentor prevention of osteoarthritis. The therapeutic agents can be deliveredin a composition, as described above, or by themselves. They can beadministered systemically, or can be targeted to a particular tissue.The therapeutic agents can be produced by a variety of means, includingchemical synthesis; recombinant production; in vivo production (e.g., atransgenic animal, such as U.S. Pat. No. 4,873,316 to Meade et al.), forexample, and can be isolated using standard means such as thosedescribed herein.

[0153] A combination of any of the above methods of treatment (e.g.,administration of non-mutant MATN3 polypeptide in conjunction withantisense therapy targeting mutant MATN3 mRNA; administration of a firstsplicing variant encoded by MATN3 in conjunction with antisense therapytargeting a second splicing encoded by MATN3), can also be used.

[0154] The invention will be further described by the followingnon-limiting examples. The teachings of all publications cited hereinare incorporated herein by reference in their entirety.

Exemplification

[0155] Identification of the MATN3 Gene with Linkage to Osteoarthritis

[0156] Genome-Wide Scan and Linkage Analysis

[0157] Selection of Patients

[0158] A genome-wide linkage scan was performed for 329 familiescontaining 1143 individuals with primary hand OA, along with 939genotyped relatives. A list of patients with OA of the hand was obtainedbased on patients' records at hospitals and health care centers inIceland. The encrypted patient list was cross-referenced with thecomprehensive Icelandic genealogy database (Gulcher and Stefansson(1998) Clin. Chem. Lab. Med. 36:523-527; and Gulcher et al. (2000) Eur.J. Hum. Genet. 8:739-742) and pedigrees with two or more affectedrelatives, related within a distance of five meioses or less wereidentified. Patients within these families, and up to three first-degreerelatives, were recruited and examined by a rheumatologist or anorthopedic surgeon. Additionally, a group of patients and theirrelatives from another on-going study of hip OA and knee OA also hadtheir hands examined for this study. Individuals were classified ashaving primary hand OA if they met either or both of the following twocriteria: 1) OA with a minimum of two nodes at the distalinterphalangeal (DIP) joints on each hand (DIP phenotype); or 2) OA ofthe thumb with squaring or dislocation of a first carpometacarpal (CMC1)joint (CMC1 phenotype). Only individuals with idiopathic OA wereincluded in the patient cohorts.

[0159] Microsatellite Markers and Maps

[0160] The framework genomewide scan used a 1000 microsatellite markerset that contained markers from the ABI Linkage Marker (version 2)screening set and the ABI Linkage Marker (version 2) intercalating set,in combination with 500 custom-made markers. All markers wereextensively tested for robustness, ease of scoring, and efficiency inmultiplex PCR. Marker positions were obtained from the genetic mapdescribed by Kong and Cox ((1997) American Journal of Human Genetics61(5):1179-1188). In the framework set, the average spacing betweenmarkers is approximately 4 cM. PCR amplifications were set up, run andpooled on Gilson Cyberlab robots. The reaction volume was 5 μl, and foreach PCR reaction 20 ng of genomic DNA was amplified in the presence of2 pmol of each primer, 0.25 U AmpliTaq Gold, 0.2 mM dNTPs and 2.5 mMMgCl₂ (buffer was supplied by manufacturer, Applera). Cycling conditionswere: 95° C. for 10 minutes, followed by 37 cycles of 94° C. for 15seconds, annealing for 30 seconds at 55° C. and 1 minute extension at72° C. The PCR products were supplemented with the internal sizestandard and the pools were separated and detected on 3700 Sequencersusing Genescan v3.0 peak calling software (Applera). Alleles wereautomatically called using DAC, an allele-calling program developed atdeCODE genetics (Fjalldal et al. (2001) Proc. Int. Joint. Conf. NeuralNetworks A1-A6), and the program DecodeGT was used to fractionate calledgenotypes according to quality and edit when necessary (Palsson et al.(1999) Genome Res. 9:1002-1012).

[0161] Statistical Methods for Linkage Analysis

[0162] Multipoint, affected-only allele-sharing methods were used toassess the evidence for linkage. All results, including LOD and NPLscores, were obtained using the program ALLEGRO (Gudbjartsson et al.(2000), Nature Genetics 25(1):12-13). The S_(pairs) scoring function(Kruglyak et al. (1996) American Journal of Human Genetics.58(6):1347-1363; and Whittemore and Halpern (1994) Biometrics50:118-127) and the exponential allele-sharing model (Kong and Cox(1997) American Journal of Human Genetics 61(5):1179-1188) were used togenerate the relevant one degree of freedom statistics. When combiningthe family scores to obtain an overall score, instead of weighting thefamilies equally, the default of GENEHUNTER (Kong and Cox (1997)American Journal of Human Genetics 61(5):1179-1188), or weighting theaffected pairs equally, a weighting scheme that is half way between thetwo in the log scale was used; the family weights are the geometricmeans of the weights of the two schemes. While not identical, thisweighting scheme tends to give similar results to that proposed by Weeksand Lange ((1988) Am. J. Hum. Genet. 42:315-326) as an extension of aweighting scheme of Hodge ((1984) Genet. Epidemiol. 1:109-122) designedfor sibships. The P value was computed two different ways and the lesssignificant one is reported. The first P value was computed based onlarge sample theory; Z_(lr)=✓(2 log_(e) (10) LOD) is distributedapproximately as a standard normal random variable under the nullhypothesis of no linkage (Kong and Cox (1997) American Journal of HumanGenetics 61(5): 1179-1188). Furthermore, because of the concern withsmall sample behavior, a second P value was computed by comparing theobserved LOD score to its complete data sampling distribution under thenull hypothesis (Gudbjartsson et al. (2000) Nature Genetics25(1):12-13). When a data set consists of more than a handful offamilies, which is the case herein, these two P values tend to be verysimilar. To ensure that the results were a true reflection of theinformation contained in the material, for a linkage result to beconsidered significant, not only was it required that the P value besmaller than 2×10⁻⁵ (Lander and Kruglyak (1995) Nature Genetics11(3):241-247), but also that the information content in the region wasat least 85%. For the families in this study, an information content of85% corresponded to a marker density of approximately one marker everycentimorgan. The information measure used has been defined previously(Nicolae (1999) Allele sharing models in gene mapping: a likelihoodapproach. PhD thesis. University of Chicago, Chicago) and implemented inALLEGRO. This measure is closely related to a classical measure ofinformation (Dempster et al. (1977) J. R. Stat. Soc. B. 39:1-38), havingthe property that it is between zero, if the marker genotypes arecompletely uninformative, and one, if the genotypes determine the exactamount of allele sharing by descent among the affected relatives.

[0163] Results of Linkage Analysis

[0164] An LOD score of 1.48 was observed on chromosome 2. In order tostudy the effects of the sub-phenotypes on the linkage results, threeadditional genome-wide scans were performed, in which individuals wereconsidered affected if they had the DIP phenotype (DIP cohort), the CMC1phenotype (CMC1 cohort), or both the DIP and the CMC1 phenotypes(DIP/CMC1 cohort). The DIP cohort scan included 944 affecteds in 274families; the CMC1 cohort scan included 558 affecteds in 204 families;the DIP/CMC1 cohort scan included 382 affecteds in 142 families. Table 1indicates the location and size of peaks on chromosome 2 with an LODscore above 1 for the primary hand OA cohort, the DIP cohort, and theCMC1 cohort. No LOD scores above 1 were observed for the DIP/CMC1 cohorton chromosome 2. TABLE 1 Cohort LOD Score Marker Location (cM) Primary1.48 D2S146  51.5 Hand OA Primary 1.14 D2S2277 160.4 Hand OA DIP 1.13D2S2324 160.8 CMC1 2.23 D2S2168 48.0

[0165] Only one location on chromosome 2, achieved an LOD score of twoor greater in at least one of the four scans (at D2S2168 for the CMC1cohort). Markers in this region were added to increase the informationcontent on allele-sharing among affected relatives.

[0166] Table 2 summarizes the finemapping linkage results for thechromosome 2 locus, indicating the peak markers, along with theirgenetic locations. Increased evidence for linkage on chromosome 2 forall cohorts was seen, with an LOD score of 4.97 between D2S175 andD2S2201 for the CMC1 cohort (p-value of 8.5×10⁻⁷). This LOD scoreremained significant even after correction for the four genome-widescans. For the CMC1 cohort, the size of the region on chromosome 2 thathas an LOD score within one of the peak LOD score is a little over 5 cMfrom D2S175 (41.9 cM) to D2S1324 (47.1 cM). TABLE 2 Cohort LOD ScorePeak Location Peak Marker(s) Primary 2.42 44.0 D2S175, Hand OA D2S2201DIP 2.44 44.0 D2S175, D2S2201 CMC1 4.97 44.0 D2S175, D2S2201 DIP/CMC14.44 44.0 D2S175, D2S2201

[0167] Screening for Mutations in the MATN3 Gene

[0168] Based on these results, and primarily on the result in the CMC1cohort, association analysis at the chromosome 2 locus is ongoing. Sixpublicly characterized genes were found to be within a 4 Mb regioncentered on the chromosome 2 peak. One of the genes, MATN3, is locatedwithin 100 Kb of the LOD score peak, and a recent publication implicatedmutations in this gene to a class of dysplasias of large joints withassociated early-onset OA (Chapman et al. (2001) Nature Genetics28(4):393-396).

[0169] To identify polymorphisms within the MATN3 gene, primers(described in detail below) were designed for PCR amplification of allknown exons and the promoter sequence of the MATN3 gene, as well as mostof the intronic sequence. DNA from 76 patients belonging to familiesscoring positive in a non-parametric-linkage analysis for the markersunder the peak of the LOD score and 18 controls were initially sequencedfor polymorphism within the gene. Both the forward and reverse strandswere sequenced on ABI prism 3700 DNA analyzer.

[0170]FIGS. 6A to 6B list all primers used for PCR amplification of DNAsequences of the MATN3 gene. Subsequent sequencing of both forward andreverse strands revealed the nucleotide variations listed in Table 3.

[0171] Results of Mutational and Association Analyses

[0172] In the initial mutational analysis carried out on the exons ofMATN3 in samples from 76 patients and 18 controls, a novel coding SNP,which showed excess in patients was identified. This mutation involves anucleotide change from cytidine to thymidine in the third exon of thegene, predicting a substitution of a threonine by a methionine residueat amino acid position 303 in the first epidermal growth factor-like(EGF) domain of the MATN3 protein. The gene for MATN3 contains 4 repeatswith homology to the EGF domains (Wagener et al. (2000) Mammalian Genome11(2):85-90). Alignnent of partial EGF amino acid sequences (FIG. 3)shows that the threonine resudue is conserved within all of the EGFdomains in this position, in human (HuEGF1-4), mouse (MouEGF1-4) andchicken (ChEGF1-4). The mutated threonine is shown in bold, andasterisks denote conserved amino acids.

[0173] The excess in patients, with a relative risk of around two,persisted after genotyping this SNP for 190 more patients and 190 morecontrols, but the frequency of the mutation was very small, only alittle over 1% in patients.

[0174] In order to more fully investigate the contribution of thismutation to hand OA risk in Iceland, the entire patient set was typedusing a fluorescent polarization method (Chen et al. (1999) Genome Res.9:492). A total of 2162 patients and 873 controls were typed for thiscoding SNP. Among the patients, 1312 of them had the CMC1 phenotype. Theresults of the mutation screening for the MATN3 gene are shown in Table3. TABLE 3 Location Base no. Polymorphism Amino acid change 5′prime37916 c/t 5′prime 38157 0/t 5′prime 38211 c/t 5′prime 38270 c/t 5′prime38367 0/ggggcggggc 5′prime 38374 g/a 5′prime 38390 a/c Exon 1 38496 c/tPro/Ser Exon 1 38527 g/t Leu/Arg Exon 1 38652 g/t Ser/Ala Intron 1 39565a/c Intron 1 41240 a/g Intron 1 41763 c/t Intron 1 42446 c/t Intron 142795 a/g Intron 1 43239 c/t Intron 1 43580 0/g Exon 2 44927 g/a Val/MetExon 2 45010 c/t Exon 2 45171 c/t Val/Ala Exon 2 45178 g/a Exon 2 45246c/t Ala/Val Exon 2 45317 a/g Glu/Lys Intron 2 45366 0/tc Intron 2 454340/tctt Intron 2 45506 a/g Intron 2 45507 c/t Intron 2 45584 c/t Intron 245588 a/g Intron 2 45661 g/t Intron 2 45808 a/g Intron 2 45852 a/gIntron 2 46055 a/t Intron 2 46104 c/t lntron 2 46168 g/t Intron 2 46404a/g Intron 2 46804 a/g Intron 2 46960 a/g Intron 2 47482 c/t Intron 247712 c/t Intron 2 47753 c/t Intron 2 47769 a/tcc Exon 3 47812 a/g Exon3 47852 a/g Val/Ile Exon 3 47864 c/g Asp/His Exon 3 47928 c/t Thr/MetExon 3 47929 a/g Intron 3 47950 a/g Intron 3 48047 c/t Intron 3 48064c/g Intron 3 48120 a/g Intron 3 48936 a/g Exon 4 49045 c/t Intron 449400 c/t Exon 5 50584 g/t Asp/Tyr Intron 5 51769 c/t Intron 5 523180/at Intron 5 52356 c/t Intron 5 52757 a/g Intron 5 52792 c/t Intron 553007 a/t Intron 5 53327 0/gt Intron 5 53482 a/c Intron 5 53828 a/g Exon6 53862 c/t Ser/Phe Exon 6 53900 a/g Ala/Thr Intron 6 54077 c/t Intron 654747 a/t Intron 6 54752 a/t Intron 6 55775 g/t Intron 6 56260 g/tIntron 7 56822 c/g Intron 7 56911 c/t Intron 7 57032 c/t Intron 7 57927c/t Exon 8 57981 c/t Arg/Cys 3′ 58045 c/t 3′ 58162 a/t 3′ 58407 a/t 3′58521 g/t 3′ 58721 c/t 3′ 58818 a/g 3′ 58878 0/tt 3′ 58883 g/t 3′ 58892c/t 3′ 59224 a/g 3′ 59426 a/g 3′ 60875 a/g 3′ 60877 a/c 3′ 60965 a/g 3′61007 c/t 3′ 61213 c/t 3′ 61256 a/g 3′ 61261 c/t

[0175] Nine of the 873 controls were heterozygous for the mutation, nonewere homozygous, whereas 43 of the 2162 patients were heterozygous, and2 were homozygous. The relative risk for OA in individuals with thethymidine residue at position 47928 was also determined, as shown inTable 4. TABLE 4 Association Analysis of Mutation at Nucleic Acid 47928Cntrl. Cohort RRisk # Affected Aff. Freq. # Controls Freq. Primary HandOA 2.12 2162 0.0109 873 0.0052 Knee OA (DIP) 2.06 1801 0.0106 873 0.0052Thumb OA 2.61 1312 0.0133 873 0.0052 (CMC1) Both Finger & 2.67  9510.0137 873 0.0052 Thumb (DIP/CMC1)

[0176] The mutation was present in 2.1 % of patients with hand OA in theIcelandic population. The estimated relative risk (RRisk) of primaryhand OA for carriers of a single copy of the mutation compared to thenon-carrier under the multiplicative model is 2.12. Both of thehomozygous carriers and 31 of the 43 patients heterozygous for themutation had the CMC1 phenotype. This led to an estimated relative riskfor the CMC1 phenotype of 2.61, which is slightly higher than that forprimary hand OA. The highest relative risk was observed for patientswith OA in both finger and thumbs (2.67). The Affected Frequency and theControl Frequency in Table 4 were also determined as described by Chenet al. ((1999) Genome Res. 9:492)).

[0177] In addition, people with the methionine mutation at position47928 have the phenotypes as shown in Table 5. TABLE 5 Phenotype # ofPeople Finger OA 30 Knee OA 36 Thumb OA 33 Hip OA 34 Back OA 17

[0178] Though this mutation alone could not account for the significantlinkage result for the CMC1 cohort, it was observed that 30 of the 45patient carriers of this mutation were in the linkage families,including both homozygous carriers. A linkage analysis for the CMC1cohort less the mutation carriers was performed, in order to assess theeffect of these carriers on the locus. The chromosome 2 peak LOD scoredropped to 3.80, which demonstrates that although these carriers have asignificant impact on the linkage, there are likely, as yet undetected,associations of hand OA to either MATN3 or other genes in the region.

[0179] Identification of a Haplotype for Increased Risk ofOsteoarthritis

[0180] By sequencing the gene and surrounding sequences, several novelnucleotide variants were identified (see Table 3). Using thesepolymorphic nucleotides and reconstructing the haplotypes, anindependent haplotype from the mutation, which carries an increased riskin patients, was detected (Table 6). TABLE 6 Polymorphism 58162 5792756822 47929 45434 45317 45178 45010 p-val aff.frq N_aff ctrl.frq N_ctrlrel risk T T G 0.00193 0.3155 718 0.2523 392 1.36576 T G A 0.002170.3152 721 0.2529 392 1.35996 T G T 0.00234 0.3134 723 0.2517 3921.3565  G A TCTT 0.00277 0.2782 724 0.2195 389 1.37035 A A T 0.003260.2771 723 0.2194 389 1.36342

[0181] In addition to the mutation at position 47928, a significantattributed risk haplotype across the gene for MATN3 was found.Polymorphism refers to the position of the SNP (Table 3), used to detectthe attributed risk haplotype, aff.frq and ctrl.frq are haplotypefrequencies in affected and controls, respectively. N_aff and N_ctr arethe numbers of affected and controls, respectively. The rel_risk andp-val are relative risk and P-value for the haplotype, respectively.This haplotype, which does not carry the thr/met mutation, is in 28-32%haplotypes from patients, but only 22-25% haplotypes from controls.

[0182] Additional SNPs in the MATN3 Gene and Surrounding Sequences

[0183] Insert and deletions are further described in FIGS. 5A-5C. Theseadditional polymorphisms around and in the MATN3 gene are likely to beassociated to the disease, either alone or as a part of a haplotype.

[0184] Matrilin-3

[0185] Matrilin-3 is a candidate for an osteoarthritis gene. It is anon-collagenous extracellular matrix protein that is one of a class of 4related proteins termed matrilins 1 through 4. All 4 matrilins areexpressed in the developing skeletal system but matrilin-3 exhibits theexpression pattern most restricted to developing cartilage, especiallythe epiphyseal cartilage. The matrilins are made up of von Willebrandfactor (VWF) A domains, EGF-like repeats, and a C-terminal alpha helicalcoiled-coil domain. Matrilin-3 has a single N-terminal VWF A domainfollowed by 4 EGF repeats and the coiled coil domains while the othermatrilins each have two VFW A domains separated by 1 to 10 EGF repeatsand then the C terminal coiled coil domain. The coiled-coil domainsmediate covalent multimer formation among the matrilins through theirheptad repeats and two cysteines. The matrilins form homomultimers andheteromultimers in almost every combination with each other inproportion to the concentration of each subunit. Matrilin-3 formsheteromultimers only with matrilin-1 and these are heterotetramers withtwo subunits of each. The VWF A domain is a collagen binding domain inother proteins and matrilin-1 has been shown to bind to Type II collagenfibrils in cartilage in a periodic pattern.

[0186] Matrilin-1 also interacts with aggrecan and may also bind tointegrin α1β1 (Makihira et al. (1999) J. Biol. Chem. 274:11417-11423).Therefore, matrilin-1 may represent the link between the collagen fibrilnetwork and the proteoglycan network as well as a connection to thechondrocytes. Indeed, mice without matrilin-1 develop normally but showultrastructural changes in fibril networks in cartilage (Huang et al.(1999) Dev. Dyn. 216:434-441). Matrilin-1 is an excellent candidate genefor OA given its interaction with the collagen and proteoglycannetworks. Several groups have analyzed association of microsatellitepolymorphism in the 3′ untranslated region of the matrilin-1 gene andhip osteoarthritis. A Dutch cohort stratified on males found asignificant association of allelic polymorphism to hip OA. In contrast,British and Argentinean cohort studies failed to replicate these resultsin their population (Strusberg et al. (2002) Clin. Exp. Rheumatol.20:543-545; Loughlin et al. (2000) Arthritis Rheum. 43:1423-1425; andMeulenbelt et al. (1997) Arthritis Rheum. 40:1760-1765).

[0187] Matrilin-3 forms heterotetramers with matrilin-1 with higheraffinity than either of the respective homomultimers, therefore it isreasonable to believe that it may serve a modulating role in thecross-linking function of matrilin-1. While matrilin-1 and matrilin-3expression patterns overlap in cartilage, matrilin-3 expression ishigher than matrilin-1 in the proliferation zone where there are mainlynon-collagenous filaments while matrilin-1 has the higher expression inthe mature zone where the collagen-proteoglycan network is moreextensive. If matrilin-1 facilitates collagen fibril formation andbinding to proteoglycan, then it is reasonable to believe thatmatrilin-3 may play an inhibitory role in normal development andmaintenance of cartilage and bone. If there is too much matrilin-3because of increased synthesis or decreased breakdown, then thesematrilin-1 roles may be impaired.

[0188] While this invention has been particularly shown and describedwith reference to preferred embodiments thereof, it will be understoodby those skilled in the art that various changes in form and details maybe made therein without departing from the spirit and scope of theinvention as defined by the appended claims.

1 132 1 137870 DNA Homo sapiens 1 gaattcctct gttttggttc ttgttgtatttctacgatct gaaaggcagc atctcccttc 60 ctcctatcca gtgctccctg gaattttttcctctaatgaa ataattagga gcggaccttg 120 gtaaacgcag tctagtttcg acattggtcttcccgttggg aaagcgaaag atccagcggt 180 aacccggttg tccgcgtttc ttggactttgttttgtttca tgcctctctt gcatctccag 240 aatcgcttgg gggcgatagc ttttgcctgagttgacgcct gaccagcttt tgtcctggtg 300 cagcctcctc cctgcgcagt aggtgcttttcgtagccgac cggctgtttt tacagagacc 360 tttattgaga gccctctcca tcctggccacctaccccggt atctcctggg ttccaagatc 420 tgtgtgattc tgtcctaacc aagttgacactgttgttgcc tggtgattca ctagtaataa 480 gtttgtgatt ttttcttttg taattcaccatttatagtat ccatgcacat taaacctaac 540 ttgtctatga cttccaaata tgggattcagtttaggtagt tttaggttat tatgattttg 600 ccgaagccgt gacaaacttc ctacatggtcgacttggcca ctttaatgac ttcgcgaatg 660 tggttgtgtg ggtgacgttg gtgatactgtgcagtttttg atacttaact gcaaaagagg 720 aagagagtta tcccagacac aatctgttttctctgtactt gctgcttctc cccactcatt 780 gacttgaaat cattagtaag catttggacaaacgccagat gagaatcatc ttaaatgagg 840 cagatactgt ttcatggccc acccccgggggactacctga ctccacatga ctgctacctg 900 ccctgttgtt ttttaccgtc ttgtttctgatgtacattta aaaaacaaca cgaatgtgta 960 ggtttgcttg tgctgagttt attttcatgtttcctgtact tttatattcg gaaataccgc 1020 cttattgaca gaactatatc acattttccctcagagactc tttcgttctg tattttaact 1080 gatttggaat actaaacata gagttaatgaccatcagctg cattatgttt ctaacttcct 1140 tttatagctt ctccattttg ctgaggggctgggtctcatt tcttttgttt gtccttagct 1200 ctgtcttcta tttccctaaa tgaataattcaaaccttgag gatcctcaag gctattactc 1260 aattccgtct tcctcacccc cagaagatgactttctttcc tatttaacta agagaatcca 1320 tcggcctggc tctctcctgc agtcttagtcttcctgtttc ttctcagatc cttccctcca 1380 ctcacagaac aaaggattcc taactcctcaagaagagaat accaccattt ctatttcaac 1440 catccgaatg taattggctg tatttcttttcagtttttct caggcagatt ttttgatgct 1500 gtgggctcag ctagaaaatg aatagaatagcaggataaag tgggaactac aaacaggaaa 1560 ctcaaaatgc agagagaaaa acttcattacgggcgcttat tcttacaaaa tagggagtga 1620 ggagtttaca tctgttccct actttaagcccttgttgtga gaagggttca catacctctg 1680 aagaccatgc tttagtttca caattttaaaaagatgccat agctgttctt actttgcata 1740 gacttctcag attagtgtaa aacattttattgtaaacttc ttaaatcgtt tgacctgtct 1800 taaattgatt aagagattta ctgaccacctgcagtgtaaa aggctttggt ggacaaacaa 1860 aaggaataaa tcacaggggc catgtgtcatttgccttgtt ttcctattac ctgctaagtg 1920 ctagactctc aaatgtttct tgtctaacacagcatctgcc cttaagatat ttatggtatc 1980 acaaacttgg agcacagata tgaaaaacatatggtacagt gatgaggagg aggagaaggt 2040 gatcagtaat atcagattct aatcagaataagtactggaa aaaggccatt ggttttgttt 2100 attatagttt tgttgatgac tttcaaaagagtagcttgaa taccatagca gtgtaaagac 2160 agtctctacc aacttgggga gacatgtaacacttatgatg caagaccacc taagccaaac 2220 tgtagtataa tacttcgaag caaagaaggcttgtccaagt agtgcatttt attctcagaa 2280 gctgctttac taagacagag aaacaatatggggatgagtg agtattaaat gtcacttccc 2340 tccctaccta ccttgtcaag gtactctcttgcttgattcc tggtgaggca ataattcagt 2400 ctgcatggga gaatatttat gtttctttttacattacgtg ggacagactt ctgagatgat 2460 gattccccct gtgtcatttg gaacttgggggagcccctct ccctcgctat cagtgactgt 2520 atctgataca gtctcattgc tttctagccacggcttctac tggtctagtg ttaaagtatc 2580 gtaacaccat atactaaaat attcattgtatggtacaaaa tagtgctgtg ggacatagag 2640 aaagccagaa ccagttatta tatgcaccggaggtaccagc cctgaagtgt gagcctttcg 2700 tattttagct agaaaagaag catacctggttaagtaacag gaaatttcag aatgagctca 2760 tgtttaaatt ctggcattcc tgaaatggttttctctatca tttattgaag aatttgaagc 2820 cagttgattt taaacaagtt tcttatgaggtataaactca aatgtttatt gaataattga 2880 aaatggactt aaaagtgtgc tgtagtagatatactttgtg attaatggtg tgcactacaa 2940 gactacattt tgagcatttg taataattttatctcttaga cttaatatat ggctaatttt 3000 tcatgagtgt ctcaaccata aacagtttgaagttctgtat ctgggagaga agtaaaaaca 3060 ggtttcctta agctctattc ctttctagccctcacctctg acttcaagta agcatagctt 3120 gtttatattt gttagtttcc ttccttaaactaaggaagtt ttgtgcctct gcctttgcat 3180 atgctattcc ttctccctga aatgttgctgtcctcccaag tcactttcta aaaattttat 3240 taaggaaatt ttcatacata caaaagtagtgaagtagtac gaaaacctcc catttattca 3300 tcacccagct tcaaccatta acaacattttgccaagtcca gataatttaa tggttagctc 3360 acatgttttc gtggtaaaac tttccttttttttttttttt gagacggagt ctcgctctgt 3420 cgcccaggcc ggactgcgga ccgcagtggcgcaatctcgg ctcactgcaa gctccgcttc 3480 ccgggttcac gccattctcc tgcctcagcctcccgagtag ctgggactac aggcgcccgc 3540 caccgcgccc ggctaatttt ttgtatttttagtagagacg gggtttcacc ttgttagcca 3600 ggatggtctc gatctcctga cctcatgatccacccgcctc ggcctcccaa agtgctggga 3660 ttacaggcgt gagccaccgc gcccggcctttttttttttt tttctttttt gagacagagt 3720 cttgctctgt caccaggctg gagtgcagtggtgcgatctc ggctcactgc aacctccgcc 3780 tcccaggttc aagcgattcc tctgccccagcctcccaaat agctgggact acaggcaccc 3840 gccaccatgt cgggctaatt ttttgtattttagtagagac ggggtttcac cgtgttggcc 3900 gggattgtct cgatctcctg acctcatgatctgcctgcct tggcctccca aagtgctggg 3960 attacaggcg tgagctacca tgcccagccaagctttcttt acttcctagc cagggggatc 4020 tgtttttctc acggtgcacc tgcaaaagtttctggatcct tcttgtatgt acttattaaa 4080 tagtactcag tacacattgg taggattgtccttttctctt atttttctct ctacagcaag 4140 atgttctcct ggagggaaga gggtgatgtctttataaggg cagtggctca cgcctgtaat 4200 cccagcattt tgggaggcca aggcaggtggattgcccagg agttcgagac cagcctgagc 4260 aacatgacga aaccccctct ctaccaaaaaaaaatatata ttatattata tatatattta 4320 tatattatat tatatatata tttatataatatatatttat atatagatat ataatatata 4380 aatatatata aaacatataa aaatatatatacacacacac atacatacat atatatatta 4440 gcggggcatg gtgtggtggt gcatgcctgtagtcccagct acaggggagg ctgaagtggg 4500 aggatcactt gagccgggga ggcagaggttgcagtgagcc tagatcgcag taactactct 4560 ccatcctggg tgacagaatg agaccctgtctcaaaataaa atgtgtgtga tggctaactt 4620 attaattaaa acatttgagg tgttcattacacatttgttg aatgaatgtc tcctccaaaa 4680 aatattgtga atactcagtt gtatctgcatgtaacagtaa acaacaacaa aaaacttttt 4740 ttttttaaat tatactgtaa gttctagggtacatgtccac agcgtgcagg tttgttgcat 4800 aggtatacat gtgccatgtt ggtttgctgcacccatcaac tcatcattta cattagttat 4860 ttctcctaat gctatccctc ccccagcccgccacccgctg acaggccctg gtgtaggtgt 4920 gtgatgttcc ccaccctgtg tccaagtgttctcattgttc agttcccacc tatgagtgag 4980 aacatgcggt gtttggtttt ctgtccttgtaatagtttgc tgagaatgat ggtttccagc 5040 ttcatccatg tccctgcaaa ggacatgaactaatcctttt ttatggctaa aaaacgtttt 5100 aaagatagtc aattttgatc tttatcagataacaggcttt gcttagttct gtagattgac 5160 cctcctgtgg gtgctttctc tgcatgggaagcccttatct ggatgcctcc ttaaaccaga 5220 cacactgttt taattattat ctcagtagatactggctctg ggcaaattac tgtgggaaaa 5280 ctgaaaaatg attttttttt taaagtctccagagaatctt ctggaatagg tgtggcagtg 5340 aaagaagaaa acaaagaatt cctctcttcaggccctgtgc ttccatggat ctgatacagt 5400 tattttgtaa caagtcaaat ctaacagatttcttattgta cacacttaca taagatgtag 5460 aagctttcaa aaacaatgca caccacctgatttgactgat taaatttaca gtactgttaa 5520 cacattgagt ggtgaatccg cagcccatgctgctaaaata ttgctgcatt ctatacctct 5580 ccatattgcc aagtttagaa gttaataatttaatcttttt tttttttttt taaagagatg 5640 gggtcttgct atgttgcctg tgctggtgtgcaatggcatg attatagcac actgcagcct 5700 tgaactgggc tcaaacaatc ctcccacctcaggcacctga ccagctggga ctacaggcac 5760 acaccactgt ccccagctta atcattctttttaaattaca tttttattaa ctttctttcc 5820 tgaatacagt gaaaggaaaa agtgaatcaggaataaatat ttgtaatagt tagtacatag 5880 ttggatattt tcagagaaat tttgtattgtaagattacag ttaattttat gtattttttg 5940 aaaagctaaa catgtaattt agaaatatcaactttaatca gagataagtt gccagggctt 6000 acatgtgagc acaaggtagg cgttcgttggtgaatatctt taagatcaaa ggatgccagg 6060 agcggtggct cgtgcctgtc atcctagcactttaggaggc caatgcagta ggatcacttg 6120 aggccaggag ttcaagacca gactggacaacatagtgaga ctccatctct acaaaaaaaa 6180 tttttttaag tcacaaggat gggttttccagtgtaatact ttaagatgta agttggggtt 6240 ctaatgaaat gatgtgatct aatggaatataattaactca aagaagattt ggaaaaagct 6300 tttaatgaag tagcattgaa atttatttcactgtttcttg gtccttctga ttggccctac 6360 gatttatttt gtttaatatt tagtggtatgacccacagtg aaattaaact tgaagtgtct 6420 ctagtactaa aagaagataa tagtggatgagcgtgtgact tatccatgga agagaagaaa 6480 agtgaagagt tcttttgttt tttaagagacagacttagta ttgttgccta ggctggtctc 6540 cagagctcag gcgatccttc ccacctcagcctcccaagta gctgggacta taggtgtgag 6600 atttttagaa atccttcatg taataaaaatatttaagaag ggaaaataga actcctaaat 6660 gtttttggca tgtcattccc ttcatttagtaattgggttt gcatgctttt cttgacaaag 6720 gaatttctgt accgcagaga gctttttcactgtgactatg ctgtgtttag caaagaagag 6780 tttgcctaat tgatttcaaa ctgatttccatacacagaac atataacgtt ttagtagata 6840 tatctgatag ttcagtaatt aattcacctagttgtattag ctcatactca tacaccacac 6900 acgctggcca aaacccattg cagcaaatgtgggcaacaaa aaaaatcagc tttcaactgg 6960 ggagagccac cttgcaaaag tgattgttcctggtaagtcc tctcaagaat tgaaagatat 7020 catgccttgc ctctgaacaa tgcaaggaaagaggcttgct gctgaacata gacagtaaag 7080 tctaaacatt ttatagcctt agataatggtttctttggga aagaccttaa aataggagtt 7140 actggggaat gtttattaat aatcacgtagtgctgagaag gaggatgtct taaaaaccag 7200 acttgtgtcc tgattctccc atctgtcagctcagcaagct taacacaagt tatatagctt 7260 tactgaatct taagtttccc catctacacataggctttgg taatacttac ttcacgggtt 7320 gttgtgagga ataagtgaag taatacacgtaaatacacat gtcataaagc atatcctcaa 7380 taaatattgg ttgattccaa atttcatattgtggcattgt ttagatcacc taccctaaaa 7440 taaaacttga tctgcttatc tagcggatgttagtgaaatg acttcagctg ataaaaggaa 7500 aatgcatttc ctgtgtgttt atcattcaaggtgaaacagg agagaagtat ataatagttt 7560 gtttaaaata ctatcgatag ctggccgacacagtggctca cacctgtaat cccagcactt 7620 tgggaggcca aggctggtgg atcacttgaggtcaggagtt tgagaccagc ctggccaacg 7680 tggcgaaacc ccacctctac taaaaatacaaaaattagcc aggcttgaat ggtgcacgcc 7740 tgtagtccca gctactcggg aggctgaggtgtgagaatgg cctgaaccca agaggtggaa 7800 gctgcagtaa gccgagatcg tgccactgcactccagcctg ggtgacagag cgaggcttgg 7860 tctcaaaaaa aaaaaaaaaa aaaaaggtcccagctacttg ggaggctgag gtgtgagaat 7920 tgcttgaacc caagaggtgg aagctgcagtgagctgagat tgcgccactg cactccagct 7980 tgggtgacag agcaaggctt ggtctcaaaaaaaaaaaaag tcccagctac cccggggact 8040 gaggcaggag aatcgcttga acccaggaggtggaggctgc agtgagccga gattgcgcta 8100 ctgcactcca gcctgggcga cagagcgagactccatctca aaaactaaat aaattaaaat 8160 actactgata ggaacttaac ccttgaaaattccagatgtt cacattttca caaagggaag 8220 gagggcaaat tctgtgaact ttactatgctagataaggaa gcctaactcc aatcctacgt 8280 gagatttctg tcttgagtcg ttaacaatactctttctgag tacttaaatg cagtgaagcc 8340 tcctgggata ccacagggat tggccaacaactacaggcac aactcatttc cttttaaaat 8400 ggatttacaa ccctgggaga ctaggaaaattcagatatta taatgggatt tcagcaggct 8460 gttcctctaa agagctaaac aattttattcagaagttgct agttaatggg aaggggcagc 8520 ctctggcagc tttgtcctca cctctgttctcttttgttag taacttagat ggaaatactg 8580 attacatgct gatcctaaat tcaggaagttgagacagtaa atctgtagtg tgtcagaatc 8640 cacatccaga aaatcttcat aggcaagaatgtaataagat gaaactcaat tacaggtata 8700 ttaaggccta tgcttgggcc tctgaatccaactgtacagg tgggagctta gcacaatcaa 8760 gtgtgaaaaa gaacttggtg ttttagggagttactctcat gtgatattca gaaaatttgt 8820 aaagccaatg caggctgggt gcggtggctcatgcctgtaa tcacagcatg ttgggaggct 8880 aaagcaggag gatcacttga ggccaagagttcaagaccag cctaggcaat atggtgagac 8940 cttgtttcta acaaaaacaa aaaacctcagtacaacttca ggttgcatta attgaaattc 9000 gttagccaga atatgagaac tgagatcccattatgttctg accagggtga ctttacctca 9060 ttttaatata tgtccatcaa actggagcatgtgtaaagag gaagacaaaa gggaaatagg 9120 tagctaggaa acatgctagt cagaggaatggcttaaagat ctgcatgtct attctagaga 9180 acacagtatt tggaggctgt tttcaggtattggagcactt atttagaagg gagagcacat 9240 tatctcaaaa ggtagattta aggccgggcgtggtggctca cgcctgtaat accaacactt 9300 tgggaggcta gggctgggtg tatcacttgaggtcaggaat tagagaccag cctggccaac 9360 atagtgaaac cccgtctcta ctaaaaatacaaaaatcacc cgcgtatggt cgtgggcacc 9420 tgtaatccca gctaccaggt ggctgaggtgggaggatcac atgagcctag gaaggggagg 9480 cttcagtgag ctgagattgc accactgcactccaccctgg gcaatagagt gagactccat 9540 atcacacaca cgcacacata cacatacacacacacacaca aaagtaagac tagtagataa 9600 gtgactttca gtatcttggt aagtctcttaagcatttgag tctagctcta gaggaaatag 9660 tgagttctct gtcacctgat gagagatgctgtggaggggt ttcgggcatc tgatgaataa 9720 atataacaga tgaacagtaa cttcttcaaatccagaaatt ctgagattgg tatatgaata 9780 aataggtagc ttcgaacatg aaccatttgtaatgattctt tagttttaat ttttttctgg 9840 tgtattggtt aagcatctta tctctcaaatggttaatatt acttatactt tataaaatat 9900 ttatttttaa attattctca tagtggtaattgcgtattgt tttatctcac aggtagtctc 9960 ccactatttt atatctttgt tacttcaagtaagacaatct taaatatgag aggacatcta 10020 gagattgtgc taagattgcg tcgtgatggtaattaatgta actgatagaa tgttatcttt 10080 tccaggtagt aaacctattg atggcaattttgctgactgt ggaagtgact catccaaact 10140 ccatgccagc tgtcaacatt cagtatgaagtcatcggtaa ttactattcg tctgagagaa 10200 tggctggtat gtgtttaata gaaaaacaaaaaaatctccc ctgctaattt ttatatacag 10260 tcctgtgttg tttaatgatg gggacacattctgagaaatg tcactgggtg atattgttgt 10320 gtgaacatca tagggtgtac ttacacaaacctagatagcg tagcctacta tacacctagg 10380 cgatgtggta gagactccta ggctaaaacctgtacagcag gttactgtac cgaatactgt 10440 aggcagttat catacagtgg caaataggtgtgtatctaaa gatggaaaaa tatagtaata 10500 ctgtaaatac actattacag tgttatggaaccctcatgat atatgcagtc cattgttgac 10560 caaaatgttg ttggcccatg tctgtctgtctccacacaca ctggttattt aggggattga 10620 ttagattatg cttaagttac atttcagaaagttaaaatgg ttacataata agggtaacaa 10680 gttaaattat atttaagtca ttcttgctgaggcaggaaag tattttgatg ggtttttcag 10740 agagattcag tgaggattac aatagagcatttctagccag aaaatgtaat ggactattaa 10800 gtttttgaga aatgtgtaaa atgtaccttgaaatcaaaat aagcatgcag gcttcataag 10860 actttttttt taatgtagct actagtagaaaggagaagtt gctatggtta caatatatgt 10920 gtccctccaa aattcacatg ctggagcttaacctccaagg tgatgatatt aagaggtagg 10980 acccttgggt ggtgattagg ccatgagggctctaccctta tgaatgggat aaatgccttt 11040 cttttttttt gagacagagt tttgcgcttgttgcccaggc tggagtgcag tggtgcgatc 11100 tcggctcagg gcaacctctg cctcctgggttcaagcgatt ctcctgcctc agcctcctga 11160 gtagctggga ttacaggcac acaccaccacacccggctaa ttttttgtat ttttagagtt 11220 actccatgtt ggtcaggttg gtctcgaactcccgacctta ggtgatccac ccgcctcagc 11280 ctcccaaagt gctgggatta caggcgtgccacagcgccca gccagcgccc ttcattttta 11340 aaatcacatt tatattattt atttgtttaaaaaaagtttt ttagagaaaa ggtcttactt 11400 tgttgtccca actggagtgc agtggtacagtaatagctca ctgtaaccca gaattcctga 11460 gctcaagtga tcctcccacc tcagctgcctgagtagctaa atttttgtgg agatcgggtc 11520 tccctgtctt taccaggctg gtcttgagctcccggcttca agggaccctc ccacttcagc 11580 ctcccaaagt gctgagatta caggcatgagccactgtgcc cagcccatag cagactcttg 11640 aaagccgatt atttgaatta caaattttgctttcaagttt gataatacct gtttaatttc 11700 atattttgtt tttacctgaa ttcttctaagtggaagtttg catgttgtag cagggaagtc 11760 ctttaaattc ttaacattta tcaatcctgtatcctctaag aaaaaagtaa agaggactat 11820 tcctagttta gtgccaaata ttgatgaggttttactgccc agtgataaat tatatataca 11880 tgctgtgcat ggtggctcat gcctgtaatcccatcacttt gggaggccga ggcaggtgaa 11940 tcacttgagg tcaggagttt gagactagcctggctaacat ggtgaaatcc tgtctctatt 12000 aaaaatacaa aaattagctg ggcttggtggcgcaggcctg caatcctagc tactctggag 12060 gctgaggcaa aagaatctca tgaacctgggaggcagaggt tgcagtgagc tgagattgtg 12120 ccactgcact ccagcctggg tgacagtgagattgtctcaa agaaaaaaaa aaagttgtat 12180 atacatttac ccaataggga atattcagcagttgttctag tcattcatca ttattagtgc 12240 cctcagtatt ttttttcttt tttcatttttaaaacatctt ttgtgcatac agggtcttgc 12300 tatattgccc aggttggtct tgaactcctggcctcaagta attttcccac cttagtctct 12360 cagagtactg agattacagg tgtgggttgccgtgcccatc ccccccagtt tcttaatctc 12420 tgcccagacg gatcacctga tgttgggagtttagaccagc ctggccaaca tggtgaaacc 12480 tcgtctctac tagaaataca gaaattagctggatgtggtg gtgcttgcct ataattccag 12540 ctacttggga ggctgaggca agagaatcacatgaacctgg gaggttgagg ttgcagtgag 12600 ccgagatcgc gccactgcac tctagcctgggcgacagagc aagactctgt ctcaggaaaa 12660 gaaagaaaaa actgtccaga gtcacagaatcaactacaga tttgtgctaa gaaattcagt 12720 aagtcaggca cggtttagaa ttgcttgagattacttggct gttttgtgaa aaaattagtg 12780 aacatggatt aatggagagt tggaacaacctgaggaagag ttgagaaacg aagaagaaag 12840 tagtgtgtgt taacatagat accagggtgcaggaagcaat gaattttggt gtttgggagg 12900 ttgggtgggg aagtgaggtt aatggcgaaatttgaatttc aggttatatt tatctagagc 12960 aaagggtaat gtatgtagca ttctgagtttaatacacttg tcttttaagc aattgccagc 13020 cattccttcc agctaaaccc tactcataagacccatcttg tgtcatttct tcatattttt 13080 tcctgatgct tcaggcaaag caaattgtttctgcctctgt tgtagcactt gacacattgg 13140 attataatga tctgtttgca catcttcagattcctgggct cataagggca cagggacctg 13200 tttctctagc agagaatgtg gttcatgtgccatcctacac ccatgtcagt gagtcgttgg 13260 agcgatttaa caaataagtg ctttgtgtgaatgtctcaag gttccttact tcatgctgag 13320 actttgattt tttttttttt tttttaccctcttggcctta gatcttttta tagccaattt 13380 tgaaaatatt taaatttaat attttaaagctagctttaag tgaagttttg ttctttttgt 13440 tgtgtaagta gcttaattag atttttgtttttgttttctt agataatgcc tgtgttcttt 13500 ttgccgtctc tgttcttatg tttataatcagttcaatgct ggtttatgga gcaatttctg 13560 taagtatact gtattttcat cttttagagtttgaccttag gggagcttag taatttccta 13620 tatgtgatac aatgttatgg ttgttattttttctccagta tcaagtgggt tggctgattc 13680 cattcttctg ttaccgactt tttgacttcgtcctcagttg cctggttgct attagttctc 13740 tcacctattt gccaagaatc aaagaatatctggatcaact agtaagtgtg tatgctcatt 13800 aaacttattt ttttttcatt ttgagtaaaatgtttacatt taaattctga atcagcctta 13860 ggtattattt ttaactactc cgcttgcttttgaaagtgtc ttaaaataat tatttctgct 13920 ctcttagcta ctgtgacaaa actggcaagatgaaagtaga gtaatttctt accatccatt 13980 tggataatgc agtattaaga gttcctagaaatggatatta tggaactagt agttcagctt 14040 ttgtttttga gcatccaaag gtacagtaatttgactgtgt atggcactct ttaagtgact 14100 agcttattgt ctttacagac atctgtagcaagtattccag tgcctactat gtttttagcc 14160 ctttgctagg cactgttggt gaatggtgtggatataaaga taactgcgaa tattacagtt 14220 tagcgagcaa actgtaatat ttggggagtcaagatacatg aagatgttac ctgtgataac 14280 aagcgggagc tgtcacagca gcagcaaataaggaacttcg attaatgtca tatggagggg 14340 ggcagggaaa actctgggct gggtggccaaggagggcgtc gtggaggtgg tcagaaggag 14400 atatgggatc taggtagatg aaaattggcaggagtatcct cagttcattt ccaaagtgct 14460 gtctttcaga acttttcttc ccactaccagatttcctgcc tagcccattt tccacaccac 14520 caaattcatc attcatttat tcagaaaacattcgctggtt gctcttatgt gcaaggcact 14580 ctgctaggtc atagagatgc atcagtggagcttacttcct ggtgaaggaa tcaggaagtt 14640 gacagttaac tgatacacca ttgtaggttactgtgggagt ggatatcaca tgggtactaa 14700 acctgggctg ggaagtgaga tttgaattgtactcaggagt ttgaacagga attaagatag 14760 agaaagtata gaagacaatg ttccagataaaagagagacc ttgaggcaag aagcagcata 14820 gtgagggcac tagaagctgg ccatggtggctggcacacag agggagggtg gggaagcctg 14880 gagaaaggtg gccaggggca gggctgttggcctggtcagc cacggtcagc ggcttctgtg 14940 tggctctctg ctcatgtcat ctccgggggagaaatttcaa gtggtgcctc cctgccttct 15000 gagtaaaacc cttagcatag cattccctcatgagctccac cttctgtgtt tttctttttg 15060 tttatagatg aagatcaaaa aaacagcttttttcccaagg agacaacctg gctggggttt 15120 ggggagtagt gtgttcttaa aattttttttttctattttt ttcagtcctt tgggatttct 15180 gtttatgttg ctttttaaat gaaaatgtaattaggccttc ttgttgacat ttccattaaa 15240 aacaagcata acttaaaaaa attaaatcagctttatagaa aagaagataa aaatttatta 15300 ttccttttag agaccctggt gttttctctcctctgttttg ttcacttagt gcctcagact 15360 ggaaggtatt tcctctgttg gttagtttttaaatctgaaa ttcttctttt ataacacctt 15420 tctccctgaa cttagctcat tgccttgtacacagtcaggg ctgattaagt atttaaatga 15480 atatagctga tgcttttcag tagaacagagtatgtctgca gtaaaactta tgtcttgcct 15540 gacattgtaa ccagtgggtg tatttttgcccctgtctgat gagagttgag acaaacgatt 15600 gacatttaga aaaataattt tccccttgtagcattaaaat ggcccagttt atctgtagaa 15660 cttacacaga actgagcatc tcctgagttaatgatgcaag tgacacaggg actgatctcc 15720 agtgccatat aggcgatagt gtttttccaaaggccagaaa gatggaaaat agtgaaaagt 15780 aaagagaaaa cgagctaaat ttagactttttggagcattg tgggttatca aatgtactct 15840 aagtgattgt tagcatgcat taatggtgataaggcttaaa atttaaaagc actttgaatt 15900 aaattctgct tatacatttt tattaaatatcacttttgac tttggtgcct ttgtattctt 15960 cagatgctct tcaggttatc agcaagtgctagtgcgatgg ctgacttgta aatgtgtgtg 16020 tgtttcttta atagcctgat tttccctacaaagatgacct cctggccttg gactccagct 16080 gcctcctgtt cattgttctt gtgttctttgccttattcat catttttaag gtacgttgct 16140 gactaaatct ttaggggtga ctgagattgacactgtgaat ctaaaccttt ttgctgcctt 16200 tcaccttaca agtcacagga gattgaggatttgtggggac agtgtttatt ctggcacatg 16260 agcagagata gcttctaggg aatggtttctgttgtgtttc caaggaatgg ttgtaactgt 16320 caggatcctt ctctgagtga aaggacctactttttatgtt tttgaaaagg ggttcatgat 16380 ttgtattgaa aatctgttaa aattcccaaggttcacaaac ccagaatttc ttctgcagca 16440 cagtgggttg acctgaattg aagctccatctctggtttct ctctgaaatg ttggcgatga 16500 aactcatgac acttaaatga gagcaagtagggaaagggcc tggcctgtgc tgggtccaca 16560 cagaggtcag ctcccttcta cacagaggatgtgtggggaa agcattctaa gtcactggca 16620 ttttagagca ggtagtctgt tgagtttctagcagatgtca actttgtttt tattcttcca 16680 ggcttatcta attaactgtg tttggaactgctataaatac atcaacaacc gaaacgtgcc 16740 ggagattgct gtgtaccctg cctttgaagcacctcctcag gttagctact gttggataga 16800 gtaaaaggtc accagtcact attgactgttccaggagaag aacctggagc ttgactttct 16860 ctttcatgct gtaacctatg gtttgggcatgaaccctggg tacttttatg aacagcttaa 16920 aatccacagg attcaacatt ttccacttggaatcctgagt acctgggtcg cttgtctcat 16980 ccagtttggc tgttagattt actcattagaggtcctcatt agtgtcatgt ttgtagggga 17040 gatgacagga aacagctgtg ttaggcccatgtagcctggg gcttctaagg atggggagtt 17100 ccattgactg aaacttgtgt tgttttcagaaaacatccat atgatccaac gggggagctg 17160 acctgtctgg ggagatgaat acagtggtttcatgggtctt ttggggaagc ctacctttca 17220 cagccctcta attgctgctg tgctaccttcccaggttttc agcatataaa atatatgtgg 17280 cccatctagg gccaggcttt cagacttggagtcacttcga ttagaatgtc actcctccac 17340 tgcataggtg tgcctgttac cctcgaagacatgatataag ttcatgatct tctgactaca 17400 aactgattct gatgaaagtc aggatccaggagctagcgtc ttattgggtg tttcctattg 17460 catggtttaa cccaactcct caacacctgaagtgtaattg gcactggcac ttcatcggtc 17520 acactggaat gagggcagag caatggcacccactgcctgc ctgtctagga atccttgtcc 17580 agaagcagga ctctctctcc ccacctccctctctccaccc tccccttcgc tagcaatctt 17640 ggtaaaattc caggttattg aggcatctacttgggaacat tttagctaac agtttcttca 17700 gctcttcttt atggttttgt ataaccctatttagggcata aagtctgaaa gctgaattat 17760 ctcacaaagg attttaaagg agaattaaatctgatacaat tttttttctc tctttagtac 17820 gttttgccaa cctatgaaat ggccgtgaaaatgcctgaaa aagaaccacc acctccttac 17880 ttacctgcct gaagaaattc tgcctttgacaataaatcct ataccagctt tttgtttgtt 17940 tatgttacag aatgctgcaa ttcagggctcttcaaacttg tttgatataa aatatgttgt 18000 cttttgttta agcatttatt ttcaaacactaaggagcttt ttgacatctg ttaaacgtct 18060 ttttgttttt ttgttaagtc ttttacattttaatagtttt tgaagacaat ctaggttaag 18120 caagagcaaa gtgccattgt ttgcctttaattggggggtg ggaagggaaa gagggtactt 18180 gccacatagt ttccttttta actgcactttctttatataa tcgtttgcat tttgttactt 18240 gctaccctga gtactttcag gaagactgacttaaatattc ggggtgagta agtagttggg 18300 tataagatct gaacttttca tctgcagaggcaagaaaaat atttgacatt gtgacttgac 18360 tgtggaagat gatggttgca tgtttctagtttgtatatgt ttccatcttt gtgataagat 18420 gatttaataa atctctttaa atacttagggttgtcattgt ttttaatccg ttactttttt 18480 ttaagataaa ccctgacatt ttccattacagtgtattcag agtgccagtg tttatttttt 18540 ttttaacttt gagaatacag ttggcccttcgtatccatgg gttctgtatc catgggttct 18600 atatccatgg agttaatcga ctgtggattgaaaatatttg gggggaaaaa tggatagttg 18660 gatctgtact gaacatgtac agactttttattcttgtcgt tgttctctaa aaaatacact 18720 ataacaattt atgtagcatt cacattatattagatattat aagtaatcta gagatcattt 18780 taaaaatatg ggaggatatg catgggtcatatgcaaatac tataccattt tattataagg 18840 gacttgagca actgtggatt tgggtatctggagggagaac ctgcaaccag tcccccatag 18900 ataccaaggg acaactgcac aatgatgcacagttaattag aaatcatttt cttaatccat 18960 taactctaaa tttgtggtga agatatttcttgaatcacag gacagaaatt aataccggtg 19020 gtttgaaaag tgattgctgt agcaatcttttttttatgtc ttattttttt tctttccaac 19080 ttcttaggtt cagagggtac atgtgcaggtttgtaacatg ggtaaattgt gttgcagggg 19140 ttttgtcagt gttttgtcat ccaggttgtgagcatagtac ctgatagcta gttttttgat 19200 cctcaccctc tactctcagg taggccctggtatctattac tcccttcttt gtgtccatgt 19260 gtacttaatg tttagctccc acttataagtgagagtattt ggatttctgt tcttgcatta 19320 atttgcctag gataatggcc tccagctccatccatgctcc tgcagaggac atgatcttgt 19380 tcttttttat gactgtagta ttccataatatatatgtacc acattttcat tatccagtcc 19440 actgttgatg ggtatttagg ttgattccatgcttttgcta ttgtgaatag tgctgtgatg 19500 aacatatgtg tacatgtgtc tttttgggagaataatttat attcctttag gtatatacct 19560 gataatagga ttgctgggtt gaatggtagaatgttttaag tttgagaaat cccttatgct 19620 gctttacaca gagatggaac taatttgcattcccaccaac agtgtgtaag tgttcccttt 19680 tctctgcaac cttgccagca tatgtttctttttacttttt aatcatagct gttctgactg 19740 gggtgagatg gatttctctg attattagcatttctctaat ggttagtgat gttgagcatt 19800 ttttcattta cttattggcc atgtgtatgtcttttgagaa gtgtctgttc atgtccttta 19860 ccaatttttt tttttttttt ttgagacagggtcctcgctc tgtcacctag gctggagtgc 19920 agtggcacga tcttggctct ctgcaacctccacctcctgg gttcaagcga ttctcatgac 19980 tcagcctcct gtgtagctgg gactacaggtgtgtgccacc acacccggct aattttcgta 20040 tttttagtag agatggggtt tcaccatgttagctaggctg gtctcgaact cctgacctca 20100 agtgttccac cttcctcagc ctcccaaagtgctgagatta cagctgtgag ccactacacc 20160 cagcctcctt tgcccacttt taatgttgttgttgttgttg ttgttttgtt ttttgcttaa 20220 gttccttata gattctgaat attagacctttgtcatatcc atagtttgca aatattttct 20280 tccattctgt aggttgtctg gttgctctgttgatagtttc tttagcagag cagagaagct 20340 ctttagttta attaggtccc atttgtcaatttttgttttt gttgcagttg cttttggtgt 20400 cttcctcatg aaatctttgc cagggcctacatccagaatg gtatttctta ggtgtccttc 20460 taggggtttt gtttgttttg ttttttcagttaggttttac atgtaagtct tgatatgggt 20520 ttggatttgt gtccccatcc gaatcttatgttgaattgta atccccagca ttggaggtgg 20580 tgcctggtgg gaggtgattg gatcatgggggtggtttctc gtggtttaac atcatccttc 20640 ttgctgctgt tctcgtgata gtgagttattgcgagatctg tgtgtgtagc acctccccct 20700 tctctctctt gctcccgctc ctgcaatgtaagatgcttgg ctcccccttt gccttctgcc 20760 acgactgaaa gctccctgat gcctccccagaagctgatgc tgccatgctt cctgtacagc 20820 ctgtggagtg agccagttaa acctctgttctttataaatt attcagtctc agctatttct 20880 ttatagcagt gcaagaacag actaatagaagtctttaatc catcttaagt tatttttata 20940 tatggtgaag gtgtctagtt ctaatctgcatttggctaac cagttatccc agcaccattt 21000 attgaatagt cctttcccca ttacttatttttgtcaactt tattgaagat cagatggctg 21060 tagtttgtat ggctttattt ctgggttctgtatttggttc cattggtctg tctgtttttg 21120 taccagtacc atgctgtttt gattactgtagtcttgtagt atagtttgaa gttggataat 21180 gtgatgcctt cagctttgtt ctttctgcttaggattgcct tggctattca ggcttttttt 21240 ggttccatat gaattttgaa atagttttttctaattctgt gaaaatgcca ttggtagttt 21300 aataggaata gcattgaatc tgtaaactgctttgggcagt atggccattt taacaatatt 21360 gattcttcct atctgtgagc atggaatctttttccatttg tttatgtcgt ctctgatttc 21420 tgtcagcagt gttttgtaat tctcattatagagatctttc accttcctgg ttagctgtat 21480 tcctagctac ttaagtcttt tgtggctattgtgaatggga ttgcattctt gatttgactc 21540 tcagcttggg tattattgtt gtatagaaatgctactgatt tttgtgtatt gattttatat 21600 cctgagactg ctgaagttta tatctaggagctttgaggca gagactatgg ggttttctaa 21660 gtttagaatc acattttctg tgaaaagagatattttgact tcctctcttc ctatttggat 21720 gctttttatt tttttcgctt gcctgattgttctcactagg acttccagta ctgtgttgaa 21780 taggagtggg gagagtgggc atccttgtcttgttccagtt ctcaagcaga atgcttccag 21840 cttttgccaa ttcagtgtaa tgttggctgtggatttgtca tagataattc ttattgtttt 21900 gaggttacgt tcctttgatg cctcatagttttttaagggt ttttgccatg aagggatgtt 21960 gaattttatt gaaagcattt tctgtgtctatcgagatgat catgtggttt tgtttttagt 22020 tctatttatg tgatgaatta catgcattgatttccatata ttgaaccaac cttggatcct 22080 aggaataaag cccacttgat cacagtgggttggctttttg atgtgttgct gggttttggt 22140 tcataaaagg cagttcccct gcacacactgttgcctatta cccagttcca aagtcgattc 22200 cacattttca ggtctcctta tttatgttcttcagggatat tggcctgaag ttttcttttt 22260 ttttgttgtg tctctgccag gttttggtatgagaatgatg ctggcctcat aggatgagtt 22320 agggaggaat tctcctcaag tttttggaatagtttctgta ggattggtac cagctcttta 22380 tatgtctggt agaatttggc tgtgaatctgtttgggacct tttccagttg gtaggagtat 22440 tattattact gattcaattt cagaactcaggctggataca gtggctcatg catgtaatcc 22500 cagcactttg ggaggccaag gcgggtggattgcttgaggc caggagttcg agacctgcct 22560 ggacaatatg gtaaaaccct gtctctactaaaaatacaag gtcaatgtga aagaaaaaat 22620 attaaaagca gctagagaga aggggcaggtcagctacatc aggttaacag cagatctttc 22680 agcagaaacc ctacaagcca gaagagattggggacctata ttcagcattc ttaaaaattc 22740 caaccaagaa tttcatatcc agccaaactaagcttcataa tcaaaaaaga aataaaatcc 22800 ttttcagtac ctaggatgca agtccactacagctcgtgtg tcctgtgtcc ctgctaagga 22860 acaagttgga gatgaggacc tcttactctcttttcccatc tatatttggc acagtgaccc 22920 acaaaaagct gagggggagg gaattaaatgttttctgaat gaaataattt ctctcccctc 22980 cacaaatttt atactgctaa tagatcattttcaatataat tgggaatata ccagtacaca 23040 tactacactt acatttaaag tttctggcaataggaaacca agagatcttt cactgataag 23100 ttgtacattt tattccctct ggtgactcacgctgtttttc agtctgtggc taatttgttc 23160 acagttatcg tggacttagc tttgttctgagcctaattgt tccactatcc cttaataggc 23220 aaaggataga tgtgctattt tcagaactcctgctttcaaa gaaggaatgt gttataaata 23280 agcccaagag ttgactgaaa gaaaaagttgtacctttgct tttctgaggt tgctttttct 23340 gtaaatgtca tgactttggt tatgagaaaataatttagaa acaagatgag tgttgccaaa 23400 taatgtggaa ttccaggcac acgagagaggcaacactggg atggtggaat gaacacaaag 23460 tttggcatca gaaggttgaa attcaattcctgtcttcacc cagcgattac actctgaccc 23520 tgggaaattt aactaggatc tgagcttcagtttttgcatc tataaaatca gaaagatgca 23580 aataactgac ctatgaactc ggagttgttcttaggatcaa gtgaggtaac agacccggaa 23640 gggttggcag accataaatt actgtgcaaatattatgagg gtgtatgaga gcaaggaatg 23700 gctggaggaa gggctggaga gggagaaaagtatttgacct ggaaaccaga aaaagagtga 23760 ctttgttccc tgtctgttag aaaggagctttgtacagtag tcccccctta tgtgtggggg 23820 atatacccta agaaccccag tgggtgcctgaaaccatgga tagtaacagt accctctata 23880 tactatgctt tttcctataa tacatacctatgataaagtt taatttataa attagagtaa 23940 gagattaaaa gcaataacta ataaaatagaacaactacaa tttttttagg ttttttttat 24000 tattattata ctttaagttt tagggtacatgtgcacaatg tgcaggttag ttacatatgt 24060 atacatgtgc catgctggtg tgctgcacccattaactcgt catttagcat taggtatatc 24120 tcccaatgct atccctcctc gctccccccaccccacaaca gtccccagag agtgattttc 24180 cccttcctgt gtccatgtgt tctcattgttcaattcccac ctatgagtga gaacatgcgg 24240 tgtttggttt tttgtccttg cgatagtttgctgagaatgg tgatttccaa tttcatccat 24300 gtccctacaa aggacatgaa ctcatccttttttatggctg catagtatcc catgatgtat 24360 atgtgccaca ttttcttaat ccagtctataattgttggac atttgggttg gtaccaagtc 24420 tttgctattg tgaatagtgc cacaataaacatacgtgtgc atgtgtcttt atagcagcat 24480 gatttatagt cctttgggta tatatccagtaatgggatgg ctgggtcaaa tggtatttct 24540 agttctagat ccttgaggaa tcgccacactgacttccaca atggttgaac tagtttacac 24600 tcccaccaac agtgtaaaag tgttcctatttctccacatc ctctccagca cctgttgttt 24660 cttggctttt taatgatcac cattctaactggtgtgagat ggtatctcat tgtggttttg 24720 atttgcattt ctctgatggc cagtgatggtgagcattttt tcatgtgttt tttggctgca 24780 taaatgtctt cttttgagaa gtgtccgttcatgtccttgg cccacttttt gatggggttg 24840 tttttttctt gtaaatttgt ttgagttcatcgtagattct ggatattagc cctttgtcag 24900 atgagtaggt tgcgaaaatt ttctcccattttgtaggttg ccttttcact ctgatggtag 24960 ttttttttgc tgtgcagaag ctctttagtttaattagatc ccatttgtca attttggctt 25020 ttgttgccac tgcttttggt gttttagacatgaagtcctt gcccatgcct atgtcctgaa 25080 tggtaatgcc taggttttct tctagggtttttatggtttt aggtctaaca tttaagtctt 25140 taatccatct tgaattaatt tttgtataaggtgtaaggaa gggatccagt ttcagctttc 25200 tacatatggc tagccagttt tcccagcaccatttattaaa tagagaatcc tttccccatt 25260 gcttgttttt ctcaggtttg tcaaagatcagatagttgta gatatgcggc gttatttctg 25320 agggctctgt tctgtagaac aactacaatttaataaaagt tatctgaatg tggtttccct 25380 ctctgtatct ctctcaaaac atcttatactgtacctacct ttcttcttgt gatctgttga 25440 tctgataacc aagatgatta cgaagtgcccaataggtggg taatacatgc aggcagcgtg 25500 gatgcactgg acaaaggagt gagtcatgtccagggcagta cagagcagga cagtgtgaga 25560 gtttatcatg ctactcagaa tggcatgcaatttaaaactt ataaattatt tttttaaagc 25620 ctatggatta tttacttctg gaattttccattcagtattt ttggactgta gtcccccaca 25680 gataactgaa actttgaaaa gtaaaaccacagataaccgg ggactagtat atctgttatc 25740 atttggttag gttgcatcag agaacttgatttatattcct ggccctccca ccaattatat 25800 gaattcagta agtaattcag attgcaaatgagattgtaca ttccttcttc ctctgacctt 25860 ctgtgactca cttttggtta aacagttaattatcttttat tattattttt tagagacagg 25920 gtctttatca cccaggctgt agtgcaatggtgcaattata gctcactgca gcctcaattt 25980 cctgtcctca agcaatcctc cctccttggtctcccaaagt gctgagatta caagtgtgag 26040 ccaccacatc cagccagttc atgatcttttaaagataaaa agataattta aaaaactaca 26100 taggtaccct cttagttacc atttttggtgttcttcattc ccttgtgtag atacatattt 26160 ccatctggta taatttttct tctttcagaaagactttaac attttttaca gtgtggttct 26220 gctggtgata agttcttttg ggttttgtatatctaaaaat atttttactc actttgggag 26280 gttaaggcgg gcagatcatg aggtcaggagtttgagacca gactgaccaa catggtgaaa 26340 ccccatctct actaaaaata caaaaagtagccgggcatgg tggtgcgggc ctgtaatccc 26400 agctactcag gaggctgagg caggagaattgcttgaaccc aggaggcgga ggttgcagtg 26460 agccaagatc gctccactgc actccagcctgggcaacaga gcaagacacc atctcaaaaa 26520 aaaaaaaaat tacctttgtt tttcaaaagctatcttccct ggatgtagaa ttctaagtgg 26580 accttttttt ggtttatttt tgttttttttctttcagcac tttaaagata ttgcttcact 26640 gtcttcttga acacattgtt tctaatgagaaatttactgt catccttgac tttctctgta 26700 tggaacattt tttttctttg ggtaatgaacattttctctt atagccctga ttttgaacaa 26760 tttgtttatg atgttcctcg atgctggctttttgtttttt cacatttgtt gcatttgagg 26820 ttcattgaga ttcttaaatc tgtaggtttatagatttcat catgtttgga aaaatttcag 26880 tcattatctc ttcaaatatt ttctctgtcctctcttctcc ctccttcagg gatcctgtct 26940 ggctggaaaa gggaagataa acttcctacagtgagcagat ttgagatagg tcttgagagt 27000 tggttaggac ttgaactggt agagatgtcaatcaaagact tcgcaactta atatgaaagg 27060 atcaagagag gaaggaggtc atcaagatttctttcatgcc tgtctggcca gaggaatagt 27120 ggaataaatg gggaataaag gacagtgagcttttttaaaa aagaggaaga tgagttcatt 27180 cttggttatg attgagcatg acgtacttttgtgaccttta aatagaggag tacaaaagcg 27240 agttacaaat gtagttttat tgctcaggagtaggttggga caagagactc tgatctggaa 27300 attagcaata tagaggtggt agctgaaacaacgctggtag ttgagactcc taacatagat 27360 ggtacacaaa gtatcaagac ctagtcatttcagcccttcc atttctagta atttcttcta 27420 gggagataat agtaattcac aaagattcatgaaaaaaggt tcagtgaagc acagcttgca 27480 attaaaaaca agtagaaact acatgtcccacaatggaaaa ttggttaagt aaatgaaata 27540 tacagtgaaa tacagtacag tcattacaaatgttgtattt gaagaatata acagaaaagt 27600 gttcactata tgttaagtac agaaagcaagttataaccct acaacatagc atggtctcat 27660 ttttaataaa aatggatgca tatataagcagagacaaaag aggtaacacc acatagctta 27720 tctttgggtg gtagtctatg gatggtctttccattctttt gcttatctac agtttctaca 27780 ttctgtatag taaagtatat gttacttttatactaagaaa aaaatcgatt ttccttttta 27840 aaagctgatt tagaaaaaag cctgtggacttccacacttc atagagcatg gaaagaagaa 27900 aaggaagtgg caaatgagag agacagtgtagaatcaggac tataggacaa cctgggatga 27960 aggggctaac actggagctc ctgccaaaaacagggtgatc taaactctac caagaggcct 28020 ctgggcttgg tgatctgagc cattatgcacttaggagagc actttcctta ggatagtgca 28080 ggggaggaag gaagagcact ttccccaggatggtacaggg gaggcagtaa gaaatgggtt 28140 aagaagtgat agctgttgag aagtgtggtgtggagagaag gaggtgggag gatagtcact 28200 cattggcccc agatgtttca tgaccccagatccagatgct cagcaagtcc cattcataac 28260 aaagacatgg ggccactgtg cagtttccttgctggtggcc atttccttgc acctgctctt 28320 actgcagata caggcctgtt gccaagatcattcgaatgag atactacaaa ctgcaggaca 28380 caaactccct cggcatagca gttgcctcctgatatggttt ggctgtgtcc ccacacaaat 28440 ctcaccttga attgtaataa tccccacatgtcaagggtgg ggccacgtag agataattga 28500 atcatggggg cagtttcccc catactgttctcatggtagt gaataagtct catgaaatgt 28560 gatggttttg taaatgggag ttcccctgcacaagctctct tgcctgctgc catgtaacat 28620 gtgcctttgc ttctcctttg cctccagccatcattatgca ggctccccag ctatgtggaa 28680 ctgtgagtcc attaaacctc tttcctttataagttaccca gtctcgggta tgcctttatt 28740 agcagcaaga gaacagacta atacagtaacttggtactgg tagaataaat tggtactggt 28800 attgatattg gtaaattgct gctgtatagatactcaaaaa tgtggaagtg actttggaac 28860 tcggtaacag gcagaggttg gaacagtttggagggctcag aagaagacaa aaaaatgtgg 28920 gaaagtttgg aacttcctag agacttgttgaatggctttg aaaaaaatgc tgatagtgat 28980 atggacaata aagtccaggc tgaggtggtctcagatggag atgaggaact tattgggaac 29040 tgtagcaaaa gtgactcttg ctatgttttagcaaggagac tggtggcatt ttgcccctgt 29100 cctagagatt tttggaattt ttaacttgagagagatgatt tagggcatct gatggaagaa 29160 atttctaagt ggcaaagtgt tcaagaggtgacctgggtgc tgctaagagt attttgtttt 29220 atgtatgcac aaagatatgg tttggaataggaacttatat ttaaaaggga agcagagcat 29280 aaaagttcag aaaatttgca gcctgatgatgcaatacaaa agaaaaaccc atttcctgag 29340 cagaaattca agctgctgaa taaatttgcataagtgatga ggagccaaat gctaaccgcc 29400 aagacaatgg ggaaaatata tccagggcatgtcagaggtc ttcatggcag cccctcccac 29460 cacaggccca gaagcctagg aggaaaaaatgtttttgtgg gcccagccca gggacttgct 29520 gctttgtgca gtctgggtac ttggtgctctgcatcccacc tgtggctaaa aggggccaac 29580 aaagagctca ggccattgct tcagagggtgcaagcccgaa gccttgttgg cttccacgtg 29640 gtgttgagcc tgcaggtgca cagaaatcaagaattgaggc ttgggaacct cctcctagat 29700 ttcagaaaat atatggaaag gcctggatgtccagacaaaa gtttgctgca ggggcagagc 29760 cctcatggaa aattcctatt agggcagtgcagaagggaaa tgtggggtag gagcccccac 29820 acagagtccc aatgggcact gcctactggagctgtgagaa gagtggcacc atccttcaga 29880 ccccagaatg gtagatccac tgacagcttgcactatgcac ctggaaaaac tgcagacact 29940 cagtgccagc ctgtgagagc aaccaagaggggggctgtac cctgcaaagc cacagcagca 30000 gagctgctca aggccatggg aatctacttcttgcatgagt ttgacctgga atgtgagaca 30060 cagagtcaaa ggagatcatt ttcgagctttaagatatgac tacccctccc caggattttg 30120 gacttgcacg gggcctctag ccctttcgtttttgtcaatt tctcccattt ggcacaggtg 30180 tatttactca ttgcctgtat cacccttgtaccttggaagt aactaagttg cttttgatct 30240 tgcagtctca taggctgaag ggacttcccttctctcagat gagactttgg aactgtggac 30300 ttttgagtta atgctgaaat gagttaaagctttaggggac tgttgggaag gcatgattgg 30360 ttttgaaatg tgagtacatg agatttgggaggggctgagg cagaatgata tgatttggct 30420 gtgtccccac ccaaatctta ccttgaattgtaataatccc catgtgtcaa ggttggtgcc 30480 aggcagagat aattgagtca tgggggtggtttcccccaca ctgttcttgt ggtggtgaat 30540 aagtctcatg agagctgatg gttttataaacgggagttcc cctgcacaag ctttcttgcc 30600 tgccaccatg taagatgtgc ctgtgcttctcctttgcctt ctgccatgat tgtgagggct 30660 ccccagtcat gtagaactgt gagtccattgaacctctttt ctttataaat tacccagtct 30720 caggtatgtc tttattagca gcatgagaacagactaatac acctccagaa actggaaatt 30780 cctgggccca gaacccaacc atctgattcagcccatatag ggtgggggcc caggaatctg 30840 tctttgataa ggcctcccgg gtgattctgacagtccagtt gcaggcccac tgcatcaggg 30900 catggggact acaggcctcc ttcccctttgtttgatttgc acccagagct ggaaaagact 30960 gaacctctga gagatatgaa aaggcttccattggaggttt ctgtaaatag cacaactgca 31020 tgttttgtca taaatcagta agtctaggatgtggaaaaaa tatatcttgg gatccatggc 31080 cttgcatgtg tcctgggaga ttttagagccaggctctgca caacccgaag tcaagaaagg 31140 gcatgactct tgggagtgac aacttgttggaagcggattc caaccatgct tcccatttgg 31200 cctgagctga gagctagcag tttccctactcgcaggacac caagcaacaa gaacaaagaa 31260 attgctccac caaggctatc tacttatgaagatgaaaagc tgttagaaag tggttaggat 31320 tagatcagcc aacgtgagtg gcctcttgaggagaagccaa cctggaatga gagatgaagg 31380 gttcttgttg aatcttctaa gagaagggccctctgagggc ccccatctat ccccatcata 31440 agcaatgtct aaataaattg tgctagatcctgttgtagtt tccaaatatt tgccattttt 31500 ccagcaaaaa gttgatatta atatttcccaactgctgcca tcaggcttgg ccacgtggct 31560 tgctccagcc aataacgtga atgaaattgatttgccacct tcttgcaggt gcatgattct 31620 accatctttt ttccttcctt tgcctcaagaccagcacatt ccagattggg cctgctccta 31680 tgtctgggtg ccagaatgaa gaagacatggaggagagctg cagccaacct atgaaggaca 31740 cataacacag caagaaataa acgtcctttgttataagaac atgagtgtgg ggccattggt 31800 taccatagca taacctagcc aaagttgactaataaatagt tttttagaat tgtaaaaaaa 31860 ttatagcaat attaaagtga tttttttaaagaaaagcaac tcttaattta ctttctagcc 31920 ttggttaaat agttttaatt tccacagatgctcacatgct aatgtgctga aacagacatt 31980 acactttagg ctgcgtctcc cccaaacttcttccctagaa ggagcctggt gttggaggag 32040 gatctcactc cactgctaga gaaggccaaggaagagatgg tctgtggagc caatgcccag 32100 cacagggcag tctgttgtca gtgaagtgaaacagagcact cgccaccact caattcctgg 32160 aaaaagcagt gccaggctgt ttctcttcctacaaacaatc tatcatgcct gttgactact 32220 ctgccaagga aaggctgaat atcagctctgttccatgatc tcccatttcc tgaagtctca 32280 gtcctaacag aggttgtgaa cctgcatgtggtaaggtaga atttctggga atttagtagc 32340 aaacaatgca agcattgcct ctgccctcttggagcctatg ctctagtgga gtctgatgga 32400 atgaatgcat ttaatacata ttaaataattacaaagtgag taaggcccac tgaagcaaca 32460 aaccagacac ttgtaataga ttgaaagggcagaggcagca gaagaaaatc tatttaatac 32520 agacgttttc aaacattagt gaatcatgaattctatttag atgtttatag ccagattttt 32580 aagaaaatga aatagaaatg aatggagttgaataaaatag gaatcattgg agtgcatcac 32640 atgtgttgtc aatattgctt tataaaactcatttctgggc caggcacggt ggctcacgcc 32700 tgtaatccca gcacttcgga aggccgaggcgggcggatca cgaggtcagg agatcgagac 32760 catcctggct aacacggtga aactccatctctactaaaaa tacaaaaaaa aaaaaaaaaa 32820 attagccagg catagtggca ggcacctgtagtcccaacta ctcaggaggc ttaggcagga 32880 gaatggtgtg aacccgggag gcggagcttgcagtgagcag agatcgcacc actgtactcc 32940 agcctgggca acagagcaag actccgtctcaaaaaaaaaa aaaaacttat ttctgttgta 33000 tatatgtatt atatatttgt gcagtaactggctcataata taaaatgtgt tgttatagtg 33060 aattgtggta aaaacaaatt tgaaaaacctaatttacaat atggcttcag agaatcaact 33120 ctggggaaga cattacagtg aactggctgacttgataatt gtttttttat ttactagtga 33180 aacttaatgc tcatgcttgt acttctaaaatgaaacaata aaaattaaaa ccaatctttt 33240 attaaaggaa gcaaaagtgt tatgagttgctcttttcaaa tattgattct actattttga 33300 atagaaatat gtcttaagtt gtaaatgttagttaataata gtaggtcatt aacagccctc 33360 ttcagaactt actggcaata atgaaatgtgcttcctgtgt ttcagaaata gtgttttttt 33420 ttctttttta gagcaagcca aactttgccttcaacctcat ctcatttgca agctactctc 33480 atatagctaa gaatttggta ttaaaaataggcacaggcct taaaacacaa gagtttagat 33540 ttagtttcca tgttttctgc ttattcacagaaatattgac tgattgattg attgagacag 33600 ggtctcactc tgtcgcccag gctggagtgcagtgctacga tcatggctta ctgtagcctc 33660 aatggcctga gctgaagcag tcctcctacctcagtctccc gagtagttgg gaccataggt 33720 gtgtgccacc acacctggct aatttttaaaaaatttttca taaagatgga gtctcactat 33780 gttgcccagg ctggtcttga actcctgggctcaagcgctc ctcccacctc ccaaagtgct 33840 gggattacag gcatgagcta ccacatccagtctagaaata tttatttaat aaaataaata 33900 ttaatgggta gtaccaatcc gtaatttgtactatcttaat ccctgtggca attaaaaaac 33960 agagatggca atacaacttg taggctattgaacatattca tttttcataa ttacttgttt 34020 ttttcaggtg ggtaaattgc aaaaaacataggtaaagagt tctatacttt ctggaaaaga 34080 cttccacctt gggattgaga cagttttgaacacaaaagaa aacaaatttt tttctccaag 34140 tacatggtct gtcatttatc agatttaagaagtcttggga catcttttga caacattttc 34200 tccaatgcat ccatggaatc aaaggacttttcaacttgtt tgggggtaga ttttttggtc 34260 taaagttgga aacatatgtc ttctttcaatagaacagtgg atgtctgcac acgtattttt 34320 caatgtttac acctacaagt ctatgcacattcaagacatt ggcaacagtg gtgttattgg 34380 caagggtggt tgggatagtt ccttttcaccatatacctgt agtatatgcc acaggttagc 34440 tgtcaactcc cattccaatg catgttttcctttgccttct tcactgtgga tcaaaaaagc 34500 taagaactta cctttcaaga cccccttgtggccaggtgtg gccatgagat gtcagcagca 34560 aatatctttt ggtcctttgc tctttggcctttggtcttcc ctcttccttc atcctggaat 34620 gccaacatga tacttggagt tccagcagccttgggaacaa aatccacaca ttaagggcag 34680 caaagcagga gacaagtggt tccttaagtagcttgcaagt cttattctac tgctgaaatc 34740 ctagttgtga gaaaaataat tctcccaagatgttgaattt attgcttgta agaagtaatg 34800 caaagttgaa agtaatccta acatttggtcggtttctttt ttcttttcaa agtttagacc 34860 aagtgcatgt attacctact ttaaaaataataaaattcta aaaacaatgt aaagtgtata 34920 tataaatctt tgttgtattt cttagatttttgaatagttc aaaaaacctc tgtttggctt 34980 ttttatgacc taagtcttta ggttatcaaaaagttattct agtctttatc ttgacaaagt 35040 agtataagat aatatgcagt ggtagagaaatatagctttt gcaatattcc caagtaaaca 35100 cctgaaagca aagttaccac ctggtccagcagccagttga gttggtgaca tgcaagaagg 35160 ccctacagcc acctgaattc ctctgtcagaagagctacct ttgttttatt tgacaatctt 35220 tggacatcag atgtaagaca accgccctggagtgtgaact ctggagatac aaacttatca 35280 gacaatttct atttcattca agatggcatgactttcatac accagcatct gaaattgtct 35340 gagatactga caacgaatgt gtagggacattcaccaggga gctgagattt ttattttgag 35400 ctctttaaaa cagtgccccc cagtggtatgcaatagaaaa aaaaaatcac aagattaata 35460 tatccacatt gttggtgcaa acagaacccattcctgctag aatgagctga aaaagaggct 35520 aagcatggtg gctcacatgt ataatcctagcactttggca agctgaggtg ggaggatcac 35580 ttgagcccag gagttcaaga tcagcctgggcaacataatt agaccctgtc tctataaaga 35640 ataaaaaaaa attagttggg cgtggtggtgcacacctgtg gtcccagcaa ctcgggaggc 35700 tgaggcagga ggattgcttg aacctggaggattgcttgag cccagaaagt tgaggttgca 35760 gtgagctctg atgacaccat tgcactccagcctgggtgac agaatgagac cctgtttata 35820 ataataataa taaagaaaca gaaaaagtatatattaaggg tattatttgt cccactacta 35880 gctcaaagta caaatggaaa agccaaccattcctcaagaa acatcactcc ccagttgaag 35940 tggaattcca aatgttataa tcaatatgtgttgactacga atgtgaatag tgcccctgtg 36000 aacttgtgca atgtacacct acaccatccatatgtagccc tggatattaa gtggttcaaa 36060 taatttctgg aaaggttcaa gaatcaagcttggaagccac acagtcatca accattccca 36120 aaattatact gcaggtttgc tatatgggagacctagagtc actgctgctt ggcatagaca 36180 ccacatatga tcgatgaaga ctgggccacttatgtacatc ctagctgcaa gagatgctgg 36240 ggaagcaagt tcttggagtc aacttggggaggtgctgact catgaagttg gaaatcccta 36300 aatttaggaa ggtatttgtg atggtgaattttaatgtgtc atcttgactg ggctaatgga 36360 tgcccagata gctattaaat attatttctgggtgtgtctg tgaggatgtt tctggaagag 36420 attagcattt gaatcggtag actgagcaaagaagatcacc ttcatctatg tgagtggagt 36480 gggcatcatc caatctgttg agggcttgaatggagcaaaa agacagtgga aggcaaatgt 36540 gttctctgct ggagctgggc caaccagcttctcctgtcct tggacatcag cattcctggt 36600 tcttgggtct ctggactcag attgggccttatgtcattgg ctctcctgct cttgggcctt 36660 caggcttgga ctggattaca ccactggctttcctgggccc ccagcttgca gacagcagac 36720 tgtgggactt ctcagccttc ataatcgcatgaaccaatcc ctcataatct caattactcc 36780 ccccaccctc tctctatatg tatgtgtatataaaatttat tggttttgtt tctctcaaga 36840 accctgacta atgtagggtt caaaggctttggggatcatt ctaaaagcaa gacgaaagac 36900 aaatgtccac cagagcttgc aaatactgtatttatcaatc cctcctcttc ttggcatagt 36960 ttctttcaaa cttcttagtg tcctcttcccattattacaa acttctcgtt tcttctaagg 37020 tccttctaaa ttggtgagtt ctcagtgggagttgagtgac cttttccctc agtgtgacta 37080 tcactgccca atgtcattga tacattatacaactccccat ccccacttcc cctcagctcc 37140 tgttaaccac cattctttct gtctctatgaatttgactac tctagattcc ccatgtaaat 37200 ggaatcatac agtatttgtc tttttgtgactagcttattt cacttaacat aatactctca 37260 aggttcatcc attttgtagc atctgccaaaatttcctccc ttttaaggct gaacagtatt 37320 ccattgtacg tagatactgc attttgcttttccattcaca caaagagaga cacttgggtt 37380 gtttccgcct cttggctatt gtgtataatgctgctaccag catggggtac aaatatgaga 37440 agtgaggttt aggatcgtgt gaggaggatcacttgtagag ggaaaatagt gaaataggac 37500 gaccagatag gctctgttgg aactcgcgcccattgtttta agacgcagct caaatgtcac 37560 ctggttttag aagtcttcct ggtctccgcaggcagtcagt atctctttcc taaatccacc 37620 acattcccgt agacagacac cacatttatgcaaatctcct tctcacagac agtgagttcc 37680 atgggggtcc cctggcagcc tccagccctgtaccatgcta actcttccgg agaggaaatg 37740 gggcctcggg agagcctttg aagcctgtgttggtacaggc tgtccttcca gctaacgggg 37800 ctctctgggg aatgtatgtt tgactttctattcccggtaa tataagttca aatgtttgtg 37860 gatatttgaa gctcggatga aagcgtttctattcatgccc ctagaatctg tgaggccttt 37920 tctttctttt ttttttttta aacaagagcccacagcaaaa cttgggttac ttggctgagg 37980 aaaggaatgt agagctagga gcagggtttcatcttcagcc ccagtcgggc aaaccagtgg 38040 cagaagggaa agcagagccc aggggtccccatcccgtacc tggggcgcag gccccgctgg 38100 ggtgggaggg agagtggagg gccgggcaggctgcagccgc ggatggctgc cccccctgcc 38160 ccggtgttac tgggaggacg gaggttgcgacagcagccga gcccacccag ttgcagccgg 38220 ctcggggctt agggcagggg cgggaggtgaccaccgcgcc gctggccgcc tcgcccagac 38280 attccgtttc tgccgctgga atgcgcggacaaggctcctt gttggccttg ggcggggttc 38340 gccgggccgg cctggcgctc gaggccccggggcggggcgg ggcccgaggc cgcgggacct 38400 ttaaatccga gcctcgcgtg ggctcctggcccccgacgga caccaccagg cccacggagc 38460 ccaccatgcc gcgcccggcc cccgcgcgccgcctcccggg actcctcctg ctgctctggc 38520 cgctgctgct gctgccctcc gccgcccccgaccccgtggc ccgcccgggc ttccggaggc 38580 tggagacccg aggtcccggg ggcagccctggacgccgccc ctctcctgcg gctcccgacg 38640 gcgcgcccgc ttccgggacc agcgagcctggccgcgcccg cggtgcaggt acaggcgggc 38700 ggcgggaggg acgcgtgagc atcgaacgaggagaagccca gggcatccac caagcgcgct 38760 tcatcccggc ctcaggccga ccgtggtgcgggggaaagag gcccctcccg gatgaatcct 38820 gtgggggaaa ctgaggtcca gagaaggggagggagatgcc caatatcact catgtgtcca 38880 ggctagagat gtgggtctcc tgaagctctcactcccctgt gattcggcca ggagcacagg 38940 gcagctgggg agcgcagggg ctgctgcccagcggctgctg ctaagctccg tcccagtcct 39000 gacctctcca ttcagtgagc aaacccttcctgagactctg ctgttagttc tcttggaatc 39060 tctccatctc catccaatat cagggcgtgatctttcctcc cctcccacag tcctcaggct 39120 gcctcagact ggaaggtaag ggatgcgggtgtgagaccag gaggagatcc gaggagccgt 39180 aggatacccc aggatcaaga cactctgggtctttggcgaa caattctatt tctctttttg 39240 gtcagcaatt gattcctgaa gcagagcaaagctcatagtg cagcgaagcg ggcaggtcgt 39300 ctgacacagc ccctgacagc acatgaagggaaactgaggc atatcaagtt gaagggactc 39360 tgtgaatcag tatcagaggc cacaataaaattcagggctc cgaaatccca agcaaggtct 39420 ctttccactg ccccagccta ccaacctcttaatatggttt acataagatt gtatcttcat 39480 ttccaaaaag cgcagcttca gtcttgctaaaaagagtctg tgccaggcca tttctgggta 39540 tgcagtctca tttggccagc cacgctgcatgcctcaggct gatttcaatt gtctatcccc 39600 ttctgcatct ttctggataa gctgagaaggagctcaaggg actctttcct gactcaaata 39660 ctcagtcccc gtatttgaag tggcagatttgaaatggcaa aagcattgct gctaaggcag 39720 gaggtaccgg ggtccacagg gctagattcagctggtggct ggttgctcca ggccgggaat 39780 tgagggaaag cctttcttct ggaagttaatcttactctag tcctgacctg cctccctctg 39840 ggatgggact ggaagaggaa ggagagattcagcaccaggt ggccatgtcc ccctaagctt 39900 tactggtctt tcattaccta gagtccgaacctgatcatga atgataaccc attaataaat 39960 gcgagattcc accctgttcc aaattatctccttttagcaa ccagaaattg ctaagcttcg 40020 ctatggattt ttatttctag caggactgtaaggactcagt ggtattatga gtatgggaac 40080 attttaggag attcagattc tttctttgttcttgggaatc aaattgctgc tatttagtaa 40140 atgggagtga gattcaaaaa caactaatgattgatgcgga gtaccagtca ggaaggaacc 40200 agatgttcac agaattttct gccaccatggtcaagcctca ggtattagtt ttctatggcc 40260 actgtaacaa attactgtga atttagtgccttaagacaac acacatttat tacctgagag 40320 ttctgtgggt cagaagtctg accctggtctcactgggcta agatgaaggt gtcggcaggg 40380 ctgtgttcct tttgggaggc tctaggggagactgcatttc ccttcccttt ccaccgcatg 40440 aaggctgctg agttccttgg cttatggcctttttcttcca ttttcagagc cagctaaagc 40500 aggttgaggc cttctcccat tgagttactcacatttcttt catctgcctc cctcttctac 40560 tttataatta cattgactca ctgaataatccaggataacc tcttgtgtag agctctttga 40620 cttaatcaca cctgcaaagt ctctgctctttgccacgtaa aagtaacatg tctctggaat 40680 taggacgtgg acgtctttgg cgggggacattattctgcat accacacccc gaatagagga 40740 ttcggtgtga ataaccaagt ggagaaggaaagaatcatgc acgtttagtt ataagaacat 40800 agccatgata tttataagga caccatttgggtgctggtga aagcactgtc aattacagag 40860 ttcatatctt catgttaccc ccaccttttctttttctaat tatacaaaaa tactttgacc 40920 catctccagg aaagattttc aacataattcttcttcctag ctctagatct atgggacttt 40980 ccaggaatga agtaaaattc tggaggtttttttttttttt ttttttttta gtatttgaag 41040 agatttattc tgagacaaac atgagtgaccatggtctgtg acacagccct caagaggtcc 41100 tgagtacatg tgctcaagat ggtcagggtgcagcttgttt ttatacaatt taaggaggta 41160 tgagacatca atcaaataca tttaagaaatacattggttt ggtcctaaaa ggtgggacaa 41220 ctcaaagccg gtggtggtga ggggcttccaggctataggt aagtttaaat attttctggt 41280 tgataattgg ttgcatttgc ctaaagacctgggatccata gaaaggaaat gttcaggtta 41340 agataaaaga ctgtaaagac caatgttcttttgaagtctt atagtggcta cccttagaga 41400 caatagatga caaatgtttc ctattcagatctttttaaaa aattattatt attttacttt 41460 aagttctggg atacatgtgc agaatgtgcaggtttgttac atagggatac atgtgcgctg 41520 gtggtttgct gcacccatca acccatcatctaggttttaa gccccacata gattaggtat 41580 ttgtcctaat gctctccctc ctcttgccccccaccccctg acaggcgctg gtgtgtgatg 41640 ttcccctccc tatgtccatg agttctcattgttcaactcc cacttatgag tgagaacatg 41700 tggtgtctgg ttttctgttc ctgtgtcagtttgctgagaa tgacggcttc cagcttcatc 41760 catgtccctg caaaggacat ggactcattcttttttatgg ctgctggagt tgtttttttt 41820 ttttttaaag aaagaaagaa ttttgattcctttttttttt ttttggcttt tagagactgg 41880 gtctcattct atcatccagg ctggagtgcagtggtgtgat tgtggctcac tgcagccttg 41940 actctctagc tttaagcgat cctcctgcctcagcccccta agcagctggg actacaggtg 42000 cctgccacca tgcctggcta atttttaaactttttataga gatgaggtct ccctatgttg 42060 cccaggctgg tctagaactc cggagctcaagtgatccttc tccctcagcc tctcgaagtg 42120 ttgggattac aggcgtgagc cactgcgcctgggtttaatt ccttttgact taactaaatg 42180 catgacatac agcatatttc agggcggtgacttaaagtca acaagcagga tgtttactga 42240 atacatgaga gaatgttgat ttttgtggaggaaaatgttg cctcagagag aggtaactgg 42300 gattcagtag caagctccca gggactgggagtcaggaaac ctggagtcca atccatgctc 42360 taccactttc tagttctcac acctccttgaacctcaggtt tctcacctgt aaaatgaata 42420 taaaaaccac ctctgcacct ccagcctcacaggttgatgt gttggtacct tgcgattcct 42480 aaagtgttac tgaaaggaag attttctttttgttaattaa atacctccaa gcagcaggct 42540 aaaaacaaga agataatttt ctgcttttaaaagaaaacta gacagagaca taaggtaggc 42600 tgttccaggt aaggctttaa tttttttctataaactgctt agtctaggca ttaattttga 42660 atcacctttt tttcctacta agtattaccttcctagagta gcatcaactt ctgcctcctt 42720 attaggaaag gaaagagcct tgagtgacacttcatgcccc aggacctcat ttccatttag 42780 ggctgcatgc gtgtgttggg tgagaagggggaatgacttg aaaaagaagt ttagggctgg 42840 gcacagtggc tcacacctat aatcccagcactctgggagg ccaaggtggg aggatcacct 42900 gaggttggga gtttgagccc agcctggccaacatagtgaa accccgtctc tatgaaaaat 42960 acaaaaatta gctgggtgtg gtggcgcacacctgtagtcc cagctactca ggaggctgag 43020 gcaggagaac tgcttgaacc caggaggtggtggttgcagt gagccaagat tgcaccaccg 43080 cactccagcc tgggtgacag aatgagactccgtctcaaaa aaaagaaaaa gaagtttaat 43140 aattggcttt ttttcttgtc tcatcctgacttgtggattt gatttttgta tcgaggagag 43200 agggtgtgca aacaggggtt gggggaaacaggaaagaaca ggaaagaata gatgaacact 43260 aaaatgtact gtgatactaa actgttaggtacttttgtgt agaaaagaga taaattcttt 43320 gaaggtggga aatctggcta tctggtttatatgcttttgt agctccaggg cttcatacat 43380 gtaggccctc aaaaaatgtt tttggtgaatgaataagtca aaagagagat gaccaagaga 43440 gttagttggg gcagtagggt agataccagatcacaatccc tgcttcttac gagacatgct 43500 catgaagctg gaagctcact aaacagatacaagattaagt tggcatccct cattctactt 43560 tttttttgag atgatgtctt gctcttattgcccaggctgg agtgcaatgg caggatctca 43620 gctcactgca acctccacct cccgggtttaagcgattctc ctgcctcagc ctcccgagta 43680 gctgggatta caggcacatg ccactacgacccgctaattt ttgtattttt agtagagaca 43740 gggtttcacc aagttggcca ggctggtctcaaactcctga cctcaggtga tcctcccgcc 43800 tcggcctccc acagtgttgg gattaccaggcgtgagccac cgcgcccggg cccctctttc 43860 tacttacttt tttttttgtt ttgttttttgagacggagtc tcgctctgtt gcccaggctg 43920 gagtgcagtg gcgtaatctc agctcactgcaagctctgtc tcccaggttc acgccattct 43980 cctgcctcag tctcccgagt agctgggaatacaggcaccc actaccacgc ccggctaatt 44040 ttttgtattt ttagtagaga cggggtttcaccgtgatggt cttgatctcc tgacctcttg 44100 atttgcccgc cttggcctcc caaagtgctgggattacagg catgagccac cacgcctggc 44160 tctactttaa attagttcag ggctccagtgagcccccaaa ttcactaaac aggaaagatg 44220 gccagatgga gggagaattc agggatactctggagcagtt ttcagcccct tggaagtgga 44280 gtaaaagagg ggggccacca gaagagcccctgagtgcctt gcctcagact accattgcct 44340 gaggcttggg cacagtggtt ggcagtgaggctgtccctga gggcccaagg ccttttacat 44400 ggttcaggga cacattgtcc aattgctccagatagccaag gtacttaaga aatgtgtaga 44460 atctattaat tcccaggaat catgagggcccttgggagca aaagcttaag cagagggatg 44520 aggttctaag tgaaatttca ggattcagccagcctgcagg tgaggtattt ggggaaactt 44580 cacacctcca ggtgaattct cactagggaattctgaagtc cttggtgctt tcaacacccc 44640 gatggtcttg gggaaggtcg caggcagtggtgatgttggg gtcaggagat gctgccttgc 44700 ctggcctcct ggaatcccag agttctctggaagtgaggat tactgcattt ctaattgtgg 44760 tctgatcatt gcttggtctt ttgcaggtgtttgcaagagc agacccttgg acctggtgtt 44820 tatcattgat agttctcgta gcgtacggcccctggaattc accaaagtga aaacttttgt 44880 ctcccggata atcgacactc tggacattgggccagccgac acgcgggtgg cagtggtgaa 44940 ctatgctagc actgtgaaga tcgagttccaactccaggcc tacacagata agcagtccct 45000 gaagcaggcc gtgggtcgaa tcacacccttgtcaacaggc accatgtcag gcctagccat 45060 ccagacagca atggacgaag ccttcacagtggaggcaggg gctcgagagc cctcttctaa 45120 catccctaag gtggccatca ttgttacagatgggaggccc caggaccagg tgaatgaggt 45180 ggcggctcgg gcccaagcat ctggtattgagctctatgct gtgggcgtgg accgggcaga 45240 catggcgtcc ctcaagatga tggccagtgagcccctagag gagcatgttt tctacgtgga 45300 gacctatggg gtcattgaga aactttcctctagattccag gaaaccttct gtggtaagtt 45360 ggtcagtctt tgcttccaac tagaaaggatgttatattca gacattgtgt acccagagaa 45420 aagagtatta ttcttttttg gtggaaatttaatgaaaaac ttaaatatat ccaatgcgtg 45480 tgtgtgtgtg cacgtgtgca tgcgcgcgtgtgtgcgacca acctaaacac actctattgg 45540 tttaggaata tagagctgca ttatatggaacttggtttgc tcataaaatc ttcatgtttg 45600 cgttagaact gtgaaggtct gcaaatatccaggataaatg accctgctgt tccctgtcag 45660 ggtcaggaaa aacaaaactc aagatgcctctcctgtcccc tctgccccac cactacattt 45720 ttttcccctg atacatattt ttttagttttaccatctcta gatctaacag aactctgttt 45780 gttctcctgc agaagacagg gagaagcgtctgtccttggg agagtcagca ggcctctatt 45840 ccaatgtcac cgtcagcccg agtgtgcatttagtgaatgc tctggttttc agactgggct 45900 gtttatctga ttactctttg ttgaagaacaggattcaggg accaggcaga ttagggtttc 45960 agctcacttg aaagaaagca agatgatcttaagaaatcat tctgttaact ctgctactcc 46020 ttccatcctt taggatggaa tttgaataaggacatgctaa ttttattgga atttttagca 46080 tgaattgacc aaatctagga ctacagatccattcaatgct ggagtttaat tttagtcact 46140 gatcagaatt atgatctgtg ggactcatgctttatttttt tatcatctgg agataatctt 46200 ttgttccctc tatgtgaata tccaagcagttgtggacaaa tttctagttc aggtgaattt 46260 ttaaacatag atatcttttt gcaatataatttatttcaca cttatttatt cagcatgcat 46320 ttattgatct ctgatgatac aaagataaaggtaatccctc cataccatta agtaacattc 46380 tgggggtggg ggtgagacaa acaaatgaaccaataattaa ttacaattat acatttcaag 46440 gagactttta atctaggtta atgtgaaacgcagccatcaa tggtttgtca ggaaaaggga 46500 gatgaagtct tgctctgggg caacgtttggcctcattgca gtcagacttg gctgggatcc 46560 ccttgtcttt cctgtcccag gacactgggctttcacaagc catcactggc tttggtggca 46620 tctacttggc attcgttggc ttccaaaaagaaagtggctt tctttctccc aggaaatcac 46680 cctcccactc catgcttgtg ggtgagagcctctccatcaa catgctggtc tgtggcaaag 46740 ttgggtttca aagctagcac ctggggttgggcttcgaagg ctataaagta acaagtggtg 46800 agtggcaatc ttttccttct ggtacaaaagacctgcttgt tctgggggaa aaacgtcctc 46860 tctatggaaa aaagactgag cggcagcttgttgtagtcaa cgggttggaa agatcattcc 46920 atgaaaaggc ccttcttggg aaaatattagccttgtcctg ttgccaaggg cagtggaggt 46980 tgggttacct gccacaggag ggccagcctccttcaacagg cctccttggt ttctcagaaa 47040 tgacttaatt cattgattca tctttcaaaagaggtcacta ggaaaaaaat atccaatgag 47100 atataaatgg ctccaggagc aataagatggagatgtagtt tagctaatgt aatctcattt 47160 attttgttct tttcttttta cctgtttgtattagcatatg agttcactat ggatttagcg 47220 tatggatgac ttcacccacc ttacatattcagaagacagt tacccgaact gtcctctacc 47280 atttgcatgt tcaaatatgt accactgcccacctgtgagt tcgttctagg cttagaccca 47340 gacttgtgtg agaattctgt ttagacataggctatagaac cttggaaatc agattctggc 47400 tacaatggga caactagctc acgtcacttcaggctgtgca tttattgttc tcccatttct 47460 tgtggtgggg aagcacaaat ctttttttcatggggaggcg gataagccaa ataaaacatc 47520 caattgcctg gcctatgtaa gtgtttggcttcagctccaa actccaaatt ccctttctcc 47580 cttgagaaag tctatatagc aagatgtgtacttttaaatt agcaacaatt tctagtttaa 47640 gagtagcatt gctaataagc cagaggcagaaaacaaataa ccattttttt atagtgaagt 47700 aaagatggac tcggaaagga gcccagagagcaatgtctat tgacatggga gacggtatct 47760 gagatgtttc ctaagtgtca gtgctgactgttgtaaactt cctctcacag cgctggaccc 47820 ctgtgtgctt ggaacacacc agtgccagcacgtctgcatc agtgatgggg aaggcaagca 47880 ccactgtgag tgtagccaag gatacaccttgaatgccgac aagaaaacgt gttcaggtga 47940 ggcctgtgta gggggccgtg actgaatcaaatacttggga acctatgctc caggttttgg 48000 actggccttg tgctttgact ctctgacccctttttaccta actgcctttt ggctggttta 48060 ctggtgttca tcagcagttt tccctgaaaaggagcttagg gagaaaagtg aaatgataga 48120 taaatcagct tatttccata ctttgtaagttttaaataaa atcgtaggag aaaaggctac 48180 taagaaagag gtccaggctg agtgcagtagctcatgccta taattcccac attttgggag 48240 gctgagatgg taggatagtt tgaggctaattcgagattag cctcgagaac atggtgagac 48300 cccatctcta caaaaaataa tttaaaaaaaaaagccaggt gtggtggtgc atgcctgtag 48360 tcccagttac ttgggaggcc gagatgggaggattgtttga gaccaggagg tcgaggctgc 48420 agtgagctat gataatgcct ctgctgcactgccacagagg gagaccttct ttcagaaaac 48480 agaacccaca cacacaaaaa ggaaaagaaacgggtccaag attcttagcc cttttgagga 48540 tttgaaaatt ttcctggatt ttatttcattacatgcaagc taccctcttc ctctctagtc 48600 atagatgctc atggaatcta gaagtagcaaagaagttaga ggggtactca ctggtctggt 48660 acatagatac atacatggag tcaatgaggaatgggaatta tatatttttt tgttttgttt 48720 ttaatcctag aaaaaagttt ttgggaggaagaaagcttag agataactta attttataga 48780 taatgaaatt gagatggaca gaagggaaaaaacttaccta tgtcacaggg tctcttagtg 48840 aaagagccaa gaccggaggg ttggttcccaactcccagct gcccctccct tgtttcatta 48900 gctctgtgtg tgtgtgtgtg tgtgtgtgtgtgtgtataac tgtaagtggc gtggcccaca 48960 attatttctg ccatgctctg catgcatttacatttgtccc aatcatcttt tgatagctct 49020 tgataggtgt gctcttaaca cccacggatgtgagcacatc tgtgtgaatg acagaagtgg 49080 ctcttatcat tgtgagtgct atgaaggttataccttgaat gaagacagga aaacttgttc 49140 aggtaagtga gggaggagtc tttactattgctacttagct ctctgcctac ctacccggtg 49200 atggaagggc aggtcacatt gactgggaagttggtgtgtt ttcctctctt gccaccttga 49260 gcttttccca tttttaagtc ttatccaaaatgttagaaat ggtaacatgt atgtatctcg 49320 gcccaaatac ataaatatgt tgctcatttccttctctgta gccagcctga acaatccttc 49380 ccaaatgcta cagacaattc agtaggttgaaatcacataa caacccacaa gttacctgtt 49440 gcttacaaag ggaaagacag taactttatagtggagaaac tggcagacat caccttaact 49500 aagtgatcaa agttaacatc atcagtgatgacacatagtg acatcatgta tgacctgaca 49560 tgctgtgctc aatactgtgg acacaacagcacacctgtag tattcttgct aaaaattcag 49620 gattatgagg aaagattaga aaaacctgagagacactctc agttcacaaa atagttttcc 49680 agtacttctc agaaacgtga aagtcataaaagacaaagaa agacttagga agtgtcccag 49740 gttggagagg acggagatgt gaaactgaatgcaatgtgga atcctggatt ggatcctgga 49800 ccagaaaaag gacattaatg gcaacatttgaataaagtct gtagattagt aaatcatatt 49860 atatcaatgt taatttctgg tttgataatcatactatggt tatgtaaatt actgtttggg 49920 gaaacagggt gaagtacagg aattttttgcactgtttttg caaccttttc ataaatctga 49980 aattatctca gatgaaaagt ttaaaagtaaaaaatagttt aaaaatatat cagtagagag 50040 gaaagtgcat tgaatgaaaa tgtgtgtcttgagtttttat gctgctctga atcaaatggt 50100 ctccctcgcg ctcctttccc tcatttaaaaaataggagag ggttgaacta caatttcttc 50160 aaactctaat tctatgtttt tctgaatctaatgtaattag ccacacaact gttggtatcc 50220 cgctgagttg ggataagtac aataagcctaaaaggaatgc tagaaagtat ttatgagata 50280 cttaggttat aaaaactcaa tactgatgatttccaaaagc aattgtggaa ggtcagagaa 50340 taattattgt atgtgcagga tttcacttaactcagttaac ttctagttag aaaaaaacag 50400 tgccaatggc tttgaatagt ttccagtaagttttcctcat atgtttccag ctgcaaataa 50460 ggtttctatc ctgggtaata aaatacttttccaccttcta tttcttttct cctgactttt 50520 tgtttcacag ctcaagataa atgtgctttgggtacccatg ggtgtcagca catttgtgtg 50580 aatgacagaa cagggtccca tcattgtgaatgctatgagg gctacactct gaatgcagat 50640 aaaaaaacat gttcaggtaa gatccactcaaaaaattctt tcatacttgg gtctttcaca 50700 tgaaaccaac ttgcaactca ccagcaaagaggtttttaaa ggtaacttaa gatcaaggtt 50760 cagacattct atcacacttt tagatgaaggcacacattgc tcagagggtg gtgagctcac 50820 tggagcaaac atagattact tcagaaaattactgggaaat aggtagggag tgggggatat 50880 gagaaaaact aactaaatgt agggtccttatccaggaaaa ttttatagct ctgtacactg 50940 aagactttta agcaaagaga aaactttagtggaagtcagt tggacagtaa ctatttggtg 51000 tgataggatg aactgatctt tttccagttaaaacttggcc tggaactttt ctgtagtgtt 51060 aatcttaaag ctggatgatt ttagtaaagaagactttgtg aatgcctgaa ttagacccac 51120 acagatcttc cagtaagttt gaggccagtcactccctggc agtttctaga cgatcaatca 51180 ttggtgaaac tcagccttat acgttggccaatgagcccag gccagtgctg ttaatcctca 51240 ttaatttgat ttaaacttac tggcaggacagggtggcatt tataaatgta aaaacactcc 51300 tgagagaaat tccaagctct caaggtgccgtttagctttg gcatattgaa tgtgttttag 51360 atttgacata tattttcctc tcccttgaggacatacagac ttcatcaaga tagacaaagg 51420 gaaatggcat gtcacccagt cccctctttttttttttttt tttttttttt aaagggaaac 51480 cagtaaaaaa ataccggaca cctactttgttcccagtatg gaactaagca ttgggagcat 51540 acaaagaagc ctaagtttct gcctctgagcaagttaaatc tgggtgaaca gagagccggc 51600 tccttatgaa agcagagact gagccagggagagggactga tggaaacaaa gggctgtaga 51660 ggctggggag gtgggcatct gcgaagggctggtggtgggc cttgaaggag ggtgaaggtt 51720 tcaatggaga caagtggaaa gtggcactggcgggggactg gcttgagctg atttgttcag 51780 ggcaggatga ctaagaaagt tggggtggagagaactcacc tatggctgca cttctcaaac 51840 tagtgacaag tgtcccagtt ggtactttaagctgccagga acattgaaga cctgggaaaa 51900 caaaccaaaa acaaatttcc atttaaactaacagatagta tggaataaag tgaagcaatc 51960 tattttaaaa aaaaacataa atcaaagaaaattttcagtt aaggcagaca acactcccag 52020 atgccagtat tccatattct tcaatacctactgtaggcca tgcaggattc taggcatttt 52080 agatacagta atgtacctta atactcacagcaacgttagg aggaaactga ttcagtaact 52140 tgcccatggt tactctgcta gcaagtagcagacccagggt ttgaacccag gtacttcctc 52200 tagtggttac ataggctctt aaccactaagccttactgtc ttaatgtaca aagcacagga 52260 cagtgaagag ggacagaaga gaaagaatactagtgggaaa gttagttata tatatatata 52320 tttttttttc ttctcttttt tgagacagggtctcattctg tcacccaggc tggagtgcgg 52380 tggtgtgatc ttggctcata cagcctctacctcccgggct caagtgattc tcccatctca 52440 gcctcctgag tagctaggac tacaggcacatgccaccaca cctggctaat ttttgtattt 52500 ttttgtagag atggggtttt gacattttgtccaagcttgt cttgaactcc taagctcaag 52560 caatacacct gccttggctt cccaaagtgctgggattaca ggtgtgagcc actgtgcctg 52620 gcctatttat atattttaaa gtaatagtgaatactgctat aaagctgggg aacataccat 52680 ggatcagatc ccttctaatg gtctgcttttccgttgctag tggtaggaag gcttgtaaat 52740 taaatgtgga gccccaaata agggccaggggctggaagca tgtccatttg ccacctgtag 52800 agtgggtact aacctggaat gagggggaaaaaaatgaaac accctgctat aaggcatttt 52860 gctgaaaaaa aaatctaaat tgtagcactgaaagggaata gccatattaa ggttggaaag 52920 gcactttatg tatagattat atagcgtctcacatttcaca agaagctatt ctataggcat 52980 tagtgagcca caaaaggttc tgcccatgaaatgacattta agaggaaaat tagagggaaa 53040 taaactattt tggttcctgc aatggcagaggaggaaccta tgtcttccag gaaggctttc 53100 tggatttcaa aggagaatta ctgttttatttgtcctaggc cagtctgaac atgggatgag 53160 tatggcactt tgctctcctc tccactcaaatttgggcaaa ctgcatttgt ccctgaaatg 53220 ttatttgtga tagatagtaa gtacttcactctctaatcat acacatcatc attactaacc 53280 ctgtatcatt tccccatctg tgcttgtgtgtgtgtgtgtg tgtgtgcatg tgtctcctgg 53340 tcagtctttt ctgttacact ttgtaagctatgaaggcagg cacttggtca atatagtgct 53400 ttttgtgcag catcaaactt agctccatgtatgattaggt ccctaaaaca ttacttttgg 53460 tgatgatgat gatgatccat gagtagctggggatgctaga agcattgata gtcatcggga 53520 aaggctggag gtaagttctg gaatatagtcaagactcagt gctgcccagt gctgtgggta 53580 gaaaaaaagg aaaggtaccc ttagatatctgtgagagata taaaggaaag gagtgtgtgt 53640 gatgtgaggc caaagggatg agaggtatgggaagagaaaa caggcttgct cctgcagcat 53700 ggggcctgtt gctgactact gactactacattctcagcac ttgtggggac caaatggaag 53760 cttctcctct caaacatttg tgtttacttttctatttaat gtgtgaatat tttctttcct 53820 gcaccacgtg cggtcagtcc gtgacaagtgtgccctaggc tctcatggtt gccagcacat 53880 ttgtgtgagt gatggggccg catcctaccactgtgattgc tatcctggct acaccttaaa 53940 tgaggacaag aaaacatgtt caggtgaggatcagcccttt aaggctattt cctttggtag 54000 gaaaattttt caggagcaag cttttttctttctgtctaag acacagttgt gattctttct 54060 ccctgtcaga attctccagt ggctcccactggtctacaga ataaaacaat gataccccta 54120 taccaggcag cctaccctcc atgatctgtttctgtcttct ctgtttttca catttttctt 54180 aataactaaa ctttctgcct aagtctgaattttttttatt gtggtaaaat acacatacaa 54240 tttaccattt caattatttt caagtgtatagttcagtggc attaagtaca tttacattgt 54300 tgggcaacca ccaccagtat ccatctccagaaatttttta tcatcccaaa ataaaagtct 54360 gtgccgatga aacaataact gcccatcacccgctcccctc agcccatggt aactacttct 54420 ctcatcctgg cttttaaaac ttttctactacatttgcccc cagattcact gcttcttcct 54480 gcatcctggt ccagccacag tgggctctttgatggtccag gagcatgccc tgcactcact 54540 cattcccacc ctcccctcaa tgcttttgcttgtcagaatt tgcttgccaa catttggttt 54600 tgttgaatta aatttttgcc aatccagtggatgttttttt catttcctgg ttactaatga 54660 ggtgaggcat cttttcctgg agagtactgccattttaaca gtctttcatt gcattaacat 54720 ggtatgtctt taatttaatt taattttattttattttagg tcttctttaa tttctttcaa 54780 aattgttttg tagttttcag tgtacaagtcttgtactttt aaaaatattt attcctaact 54840 attttattcc ttctggtgct attataagcaccatttctcc atttctcaac tttcaatttt 54900 gctacaaccc aggtttgaga aatggagtttgagatccatt tctcaacttt caatttttct 54960 acaatccagg tttacaattt tctatgaagtgttgtaatta agactatccc actgaaccca 55020 tgccctgcta gagcttagag actatcaggcacgacagaca ctgaacaagt attatatata 55080 cccatcttat gggtacaggg ataatggacatgtagttgat cttcaataga tacttcctgg 55140 tttattgact gattaactaa gatgggatttggactggctg tccctaaaag gttcaatagg 55200 attcagacag gtggatagat gatagattatatttaatgaa tttatgatct cccatcccta 55260 tattggatct ttgtaaaatg ttaagagtaaaaatgttgca gcaaaggacc atttcctgtc 55320 ctcaggcaat agaattactc cattggaagatgatacacca gtcttcagac ctgggaagtt 55380 actcttattt ctattcacaa ctttaggaggagccctgata ctgaatagga atagctgcct 55440 gactgaattc cataaagaaa taggagttacttttatcacc tctcatattt ggttaaatgt 55500 caatacaaac gtgaattttc agttccaaagtatttattgg taagaactag gactcttact 55560 tcaaaccatt aaatagaata acagtcctctagaggaaatc aggacttaac ctggttttta 55620 tccttttttt ttgttttgtt ttttgagacgacatttcaat ctgtcaccca ggctggagtg 55680 cagtggtgcg atctcagctc actgcaacctctgcctcccg ggttcaagca atcctcctgc 55740 ctcagtctcc cgagtagctg ggattacaggcatgggccac cacgcccagc taatttttgt 55800 gtctttatta gagacggggt ttcactatgttgtccaggct ggtcttgaac tcctgacttc 55860 aagcgttcca cccaccttgg cctcccaaagtcctgggatt acaggtgtga gccactgcgc 55920 ctggccctag ttttcatctt tggtatattttagccataca aatagaggac tattgtccta 55980 gttttatttt tatttattta tttttaagagacagggtctt actctgttac ccaggctgga 56040 gtgcagtggt gtgatcatgg ctcactgcagccttgaactc ctgggctcaa gtgatcctcc 56100 tgcctcagcc tctcgagtag ctaggattacaggtgtgtgc cactatgcct ggctaactat 56160 tttacttttt tgtagagaca gggccttgcttcgttgccca gggtggtctt gaactcctgg 56220 cctcaagtga tccttctgcc tcagcctccaaaagtgctgg cattatagga ataaggcact 56280 gcaccaggcc cattttcctt tcttaattacacagtattac ataaacattg tcttatttct 56340 atatccctgg cctcccccta taccaaccatgtttttctaa atttacttta tatcatgaat 56400 aatttctggc aatcacaaaa gtacagagaataaaataagg aaccctcact taagcatcac 56460 tcagctttga caattataaa cattttccccaatgtatcct aagatgaatt tttctgtgat 56520 aagtattttc tcgtcttttt gtttgttcatctatcccttt acctcttgga aaatgcactt 56580 aggtaagtct taacggtgga aatattcaactgtggatttc ctgatttgtt atgattccta 56640 tatagctatt cctagatagt atatttgtgtactgtcattg tcttcagcca ctgaggaagc 56700 acgaagactt gtttccactg aagatgcttgtggatgtgaa gctacactgg cattccagga 56760 caaggtcagc tcgtatcttc aaagactgaacactaaacat atcctttctg gaaggttttc 56820 tccttctgcc tggagccaac caaggtactacgatcgagcc aaacacttta agacattcca 56880 caggcaagtg acttatttgt taggttttaatatgagcagt ggtaatatac aactttaaaa 56940 aatttagagt tcttttctaa tggtttaatagactgtttca cccattcaat atttacatgt 57000 aaaagctgcc tattttgtcc cctagtgcttttccataact gtatatgctc ctaatgatgc 57060 tcttacataa cagatggaac atttatgctaatgactcctt aatatggctt tcttagttca 57120 gaatctgaac atgtacaaag caagtcagctgattaatggg gtgtgatatc tcagattaca 57180 cacagttgta ccctttgact agccaagtggtcaaaaagca gattttaaaa tgtaacatga 57240 tacaatctca gaaactttca aaatcagttctttcttcaaa ggagtaacta gggaaaagtt 57300 tctcagaaca gttcaagtta acaactctgttcctatgtac cccagaggag tggaatttag 57360 attgtggtag gtacaatcta ggcgtatgtagagactttta agggaggtac agataattgg 57420 aaagaatcaa tttccaaatc cttcacttgcttaagacact acttggaatc agctggcatt 57480 ttcttttccc gtctctcatt tccatatcaccattctccta acttcacaca agaaaggcat 57540 acttctcatt taccctagat ctactaaggtgcatttgtta gaacagcaga atcttgagat 57600 agccaaacaa agggacaatc caactatccctttaatcaat acttcataaa cactgcttcc 57660 cccatccctc acactaagat gctactccaattacatttta tatgaacaaa aacatctgaa 57720 gaccactgca ggggtctact tttattgtgctctctgaaga acaggatttt tgacttttag 57780 actgaattca acttttggtg agaaaacagttttgctcttt tttttcactt ctcataggac 57840 ataattattt tatcttgtga agtgtttgaaagtgtgttta tattttctaa tgtgtaaaaa 57900 taaatggtgt tggccttaac aacttgccacttgatgacat tttggagaag ttgaaaataa 57960 atgaatatgg acaaatacat cgttaaattgctccaatttc tcacctgaaa atgtggacag 58020 cttggtgtac ttaatactca tgcattcttttgcacacctg ttattgccaa tgttcctgct 58080 aataatttgc cattatctgt attaatgcttgaatattact ggataaattg tatgaagatc 58140 ttctgcagaa tcagcatgat tcttccaaggaaatacatat gcagatactt attaagagca 58200 aactttagtg tctctaagtt atgactgtgaaatgattggt aggaaataga atgaaaagtt 58260 tagtgtttct ttatctacta attgagccatttaattttta aatgtttata ttagataacc 58320 atattcacaa tggaaacttt aggtctagtttcttttgata gtatttataa tataaatcaa 58380 tcttattact gagagtgcaa attgtacaaggtatttacac atacaacttc atataactga 58440 gatgaatgta attttgaact gtttaacactttttgttttt tgcttatttt gttggagtat 58500 tattgaagat gtgatcaata gattgtaatacacatatcta aaaatagtta acacagatca 58560 agtgaacatt acattgccat ttttaattcattctggtctt tgaaagaaat gtactactaa 58620 agagcactag ttgtgaattt agggtgttaaactttttacc aagtacaaaa atcccaaatt 58680 cactttatta ttttgcttca ggatccaagtgacaaagtta tatatttata aaattgctat 58740 aaatcgacaa aatctaatgt tgtctttttaatgttagtga tccacctgcc tcagcctccc 58800 aaagtgctgg gattacaggc ttgaaagtctaacttttttt tacttatata tttgatacat 58860 ataattcttt tggctttgaa acttgcaactttgagaacaa aacagtcctt taaattttgc 58920 actgctcaat tctgtttttc gtttgcattgtctttaatat aataaaagtt attaccttta 58980 catattatca tgtctatttt tgatgactcatcaattttgt ctattaaaga tatttcttta 59040 aattatactg aattaacagt tattttgatattcaacagtt tcttatcaaa agacagaagt 59100 ttgattctcc tcctatagtt taatctcacactcctgaggc taaaagtgta agttcaggtt 59160 caaaagaaag gcttcaatat aaaattatctaaatttggat aaaataatga tgagctttac 59220 attaatataa gctcctgaga agcaaatatatcctacagat gtaaatataa aatataaact 59280 ttaaggaagg aacaaaaact aaatataccaaggaaatata taaagcatct cttacttttc 59340 aaaaattttg aaagggtatg acatagagctacagtagtat tttccagact tttttgagcc 59400 caaaatcagt aatacatatt aatataccacagagaaatgg tggtatagaa ggaataaatg 59460 agagttcatg tgcaagaagg gaaagataaagagaggtgag ttttcaggtg tcaccatcag 59520 taactagctt caactaatta tgtcttgaccagtttattgg aaactcctgg gttagataat 59580 actaatttgt ctcatgcgtc cagtatagggagcctatgga caggggccaa tgtatcttaa 59640 acagacacac aaacaatgga aagcaaacattatctgaatg ggattgggtt aagtttggaa 59700 agaattcctc atggagactg atttgtgctggtttggctag gcacaactgg cctgttaaga 59760 gggtaggacc atatataaag tggcttgggaagcagagccg gatggaatgt tgccctgcag 59820 aaagcatatt ggcttacttg aggtgggggctccctagtaa aaacatttag gattgaggga 59880 agatctgatt ttggatggag tttagatttgctaggaaagg cactagggac tattgtcaat 59940 tctataatag attgtgctgg gtgagacaaatcacccaggt caaaggattg gaggattttt 60000 attttatttt tgtctgagat gggaaaagactccaagtggg gaaattaact actgatgcat 60060 aaattacatg ctagattatg gtaaaagggaaaaccagaaa taaaaacata ggtgagacca 60120 aataatttca tgagtggttc tacttggatttctgccctat gccttagact ttgaggcttt 60180 gtaactctca agcattccaa aaggacaaaggagaatgaag aaagaaggta agaaaaagaa 60240 gagaatatgt aacagtcgac tctagacgagaaaacaccaa acgctcaaat atttgtaaaa 60300 tgggaggcca tgccccaata gcaattgtacgctgtaatta cccaacgttt acagaaattc 60360 ctttaaaagg accctcgggg ttgcggtaataattaaactg ttaaaaaata tcaaaacaag 60420 cggaataaac aaaatcttct attcctaaattgctctagag gttctccaat ccccaaagac 60480 acagagaagg gttgtaagat ctcagtacgcacatttgcac tttcatctgg ccagtggcgc 60540 cagcctatca aatacagtaa atggcttttccctttcaaga atgttggaac tttggaactt 60600 acaaaatact aaataatttg cccttattcgaacgggcgcc tctcctctta ggtttatagt 60660 aatgccacct gaggcgcggc tccatagcgatcatcacagg tgtggcggga gggagggcgg 60720 cagtgaccac gagacggggc gcagagggcgagcgcacggc tggcagtgac aggtcgcgac 60780 ccagctcctt gcggagagca gaaaggcccagacgccaggg aacaggacta gcccagctac 60840 cgcctcgcgc ctcgcgccct gcgcccacaaacccatcgac gtccgcgcag gcgcactacg 60900 ccggtgcccg gtgcgagggc gtcaccggaagtgtcccgcg gcgccccgga tgcgaccggg 60960 caacagcggt tgccagggcg acgggagctttccggagctg ctggtactcc cgattggaga 61020 cgtagaaccg ttacttgtcg agggccttagcggccgccgt gaccctctcg gggatcccac 61080 gatgttcttc tacctgagca agaaagtgagtttcctgggg ggcgtcctcg tttctccgcc 61140 attcccgcag cctgcggcga ttcccgctgccttcccgaga agccaggctc cttcacccgg 61200 tcctggtcgt ggcttcgcca ttccgcagccatccttcccg ccgggtccag cctccgacct 61260 cttcctgcgg ttcccagagc ctcagtctttggccgagctt gctgccctct cgcagcactc 61320 tcccattcct gcccctggct aggtccttttgggtggcttg aggataagtc gccacgtgct 61380 accctatgtt tcctcctcca cttgttgctgatccacgtga tttaaactct ttggatcacc 61440 acgagccatt gggcaagaga aaagaaataacaataggata gcctaagttg taggtaaaaa 61500 ggaagaggga ctggtggtat ggatttataccctctccccg tcctcccttt aaaaaatcca 61560 tccggtgaga ctctggcgtt ggcttcctccgcagtcacta tccttctctt ttgtgcactt 61620 ctttccctct ttcccaaatg ctagagcccgggtcagttca ggggaccatg cccattttca 61680 caacagctca ttttttttct gcaaggccgttcgtttttct tcttggaaga ttttcttcag 61740 ctttcatctc ctgcagactt acttcccagctccttcgtga aaagttgcta gtgcggctgt 61800 ggttgctttt tcagatttcc attcccaataacgtgaagct gcagtgtgta tcctggaaca 61860 aggaacaagg gttcatagca tgcggtggtgaagatggatt actgaaagtt ttgaaattag 61920 agacgcagac aggtaaatga atgcaagcccacatgtttgg tagtaaattg gctaatgtta 61980 tttgttcagt gcggtctact catgttatctgtcaatatgc agttctgatt attttaaagg 62040 gaaaactacc aaaacatttt atggtgtttttctggagtat atttgcaata aagttcagag 62100 aatgttgaat gtaaacatcg gatggtttttgctgagttat taatggaaaa tgaaaaataa 62160 tgaagaatga aaataagtca ctgtgttatgctatgtctaa ttattgtact atattattag 62220 gagtctaaaa gaaatcctgt gagtcaacatttcactaaga aaatctggca gattgtttac 62280 aggtctgtgt catcctttct gcttatcagatgcattaatg gttattcacc tagtgtccaa 62340 gaacagaaaa tgggatacgt tttacatttctattagtagg tctcttgtct ttgctgcttg 62400 atttctttct ttatctagtt ctctggaatcacagcaaaca gcttcccact tcttagcccc 62460 cctagattga agacctagat aaggtaattggagttaaatc tcagctcttc cacaagacta 62520 ctgaatgact ttcaacaagt tagctgaggatcttactact ttagtttagt cttccataaa 62580 atgaaaatgt agggggttaa actgcattacatttttggat tccttacaac atagcttctc 62640 aaactgtatt aggtataaga atcactggaagaggttgtta agatggattt catgctctac 62700 ccccacacaa tagacccggg atggggcctgagaatttcca tttctagtga agctgagttg 62760 ctggtcctca gaccgcactt tgaatagcattaccttacaa ctttgaatgc ggtatgaata 62820 ataaatactt attgacgatt ttccatgtagtggatgttta accttcacaa tcttatttga 62880 cttcacaata atcttgggca gtaggtattctaggtgagaa aattgaggct ctgagctgtt 62940 aactgcccta ggtcacgcag ccagtaagtggcggaacaag gatgtgaaac taagtttgtc 63000 tcacattcaa gtccttgttt ttaaccactacacaaagggc tagtggacag cctaagtaaa 63060 gaagagtacc aattaaagtt tggcttatgctgttttatag ttcaatctgt cagtgcttta 63120 gttttttaaa actgtgttaa aagaaagtttttctctgttt cttaaatgtg tatatagata 63180 tatatatacg tgtgtatata tatatatgtgtgtctgtata tacatacttt tgcttacaat 63240 atttggcaag tttttttttt tttttttttttttgagacag agtttcgctc tgttgcccag 63300 gctggagtgc agtggctgga tctcggctcattgcaagctc cgcctcctgg gttcatgcca 63360 ttctcctacc tcagcctccc aagtagctgggactacaggt gcccaccacc atgcctggct 63420 aatttttttg tatttttagt agagacagggtttcaccgtg ttagccagga tggtcttgat 63480 ctcctgacct cgtgatctac ccacctcggcctccctaagt gctgggatta caggtgtgag 63540 ccaccgtgcc tggcctgcaa gttttaaattacattaagaa gtgcattcat tttgtttaac 63600 attgaaggct tttttaaaaa caaacttctttttctgctct gcccttttga tgagctgtga 63660 ctggagtatt ctttgaaatt agtgaacaagatgggaaatc aattcttggg aggtttggaa 63720 aatggagtaa agtatttgtt ttggaactgaatgattatga attgtgctga aagaaataaa 63780 tatgaggaat atggctgtga gagtctaacaagatcagtat attatattta atacaactag 63840 tgttgattca gcagatccgt ccagtaacaatcatagatat tgcatgcctg ctttctatga 63900 agcactgtgt tgtgctaaag atgcaacatgtgaagtataa tgtgttgtag tttgggtaca 63960 gcaacataat acagcaatcc tctacgtccataaatcatct tctatggcac cattgttaat 64020 gtctatttag tatttcatta tattggtattaatatatcat aatttgctta accaatttta 64080 gtattggaca tttaggtgtc tccacatttttactattaaa gcaactgtat ctttgtatcc 64140 aaatctttgc ataaatagtt aatactttctacgataaatc tctagaaatg gcattcactg 64200 gatcgaagag tgtgtacatt tatacaagttcctaatatgt gtacacaaat taccctccag 64260 aaagtttgca ccacattgca tttccagttttcagttccct gctctagaag gaaaggaatt 64320 agaaccagca atccatcttt ttacatcaggtttacttctc tcacaggcca gcttccagtg 64380 ttttgctgga gcccatctcc cttcaccttctcaggtacct ttctgccaat ttatcctctc 64440 tctctccttt caatatcagt cctctctccttttctctccc tcaccagtga ctttcatgtt 64500 ctaaatctaa tggacagctt ttattgactcatcaggattt gacttgaaac attatttttc 64560 cagacaagag aattcagaat gtttaagttattgttatatt tagacgtact acttttctct 64620 ccttctctgg tcagtcctca gtctctttgctagatttccc tcttctatta gaattcctga 64680 gaacttgctt ctgggctctc tattcacttgctctatttat tctttcattc tcatgccctt 64740 aaatgccatc tgtggtatga ggtgaagagccccacattta tatctccagc ccagtcctga 64800 tttttcagac gtgggtgtct ccactggagtctttcacagg tatctcctat ttactgtgtt 64860 caaaaccgaa ctcttgatat tcttccaaatctctttcaat tctctccatt tcagcaaata 64920 tctccatttc actcatttgc tcaagctggaaatcttctct tttcctccgt taacaacatc 64980 tgcatcagca atcctgttgc ttttgtctccaaagtgtatg ttgaacccat cctttccttt 65040 gtctcccctg ctccctctat agctccagctaccatcatcc ctcacctggt ctattgtgag 65100 aatcagtttc cctccttcca ttgtctctggcttcctctta gtcgttcttg acactctagc 65160 cagtgtcagg cttttttttt tttttttttttttttttccc gagatggggt ctcactctgt 65220 tgcccaggct ggagtgcagt ggtgcagtcttggctcactg caacttccac ttcccgggtt 65280 caagcaattc tcctgcctca gcctcctgagtagctgggat tacaggtgcg tgccaccacg 65340 cccagctaat tttttgtatt tttttttttttttttagtag agacggggtt tcaccgtgtt 65400 agccaggatg gtcgtgatct cctgacttcaagtgactcgc ccgcctgggc ctcccaaaat 65460 gctggggtta caggcgtgag ccactgcgcccggccctttt tttttttttt ttttttaaag 65520 ggaccagatg atgtcatttc cttgccaaaaggtatttaaa gggctgacag cctgggttgg 65580 agctgcttgt ggtagggttt gaggatgatggtcctgggtt ggagctgctt gtggtagggt 65640 ttgaggatga tggtcctggg ttggagctgcttgtggtagg gtttgaggat gatggtcctg 65700 ggttggagct gcttgtggta gggtttgaggatgacggtcc tgggttggag ttgctggtgg 65760 tagggtttga ggatgatggc cctgggttggagctgcttgt ggtagggttt gaggatgatg 65820 gccctgacta ggattgaggt ggcatttcttgctttttctg ctacgactgc caaggcctcc 65880 accctcatcc tcatgagagg aaatagagagccctggaagc caacatttgc atcagagaaa 65940 aagtatatag ggccatggaa gccaactgcagctgactgcg cagtcctccg gtacctgcag 66000 gcagtgacat tgcctcaacc ttccaatatgcccaggacaa caaaattgac cagaaatctc 66060 aggtaaggtg tccttagcta gacccagggatgcatacaga gtcctttgca gtcagtcatt 66120 cacacagaga agcaaggcaa ccagatgatcctgccagaga atctggcgaa tatggtacca 66180 tctctttctc agaattgtat tatccttatgtgttgcaaaa cagtcttctg tatattttga 66240 ttctaggttt caagcttatt ttgcagcatgtaacaggaat ttctgttgaa attgggctgc 66300 tgacaacttt tatgaatgca aacaaaagcattggcagtca cgcttttcta agagaaagat 66360 tctcaaagat ttagtgtgct tggttattggtgttattagc atgatcttct gttcttctat 66420 gtcatacctt ttgttttcag ttgctttattacaccttaat tttttaaaag ccttctttgg 66480 actttttgat gtacttcgga ttgttggaataactgatttc attctgaaat tcctttacat 66540 gagtttaaaa tgctttattt tattggtgacttcttcgatt atgcctttta aatccaagga 66600 ttactgttat gtgcttttag aagatttatgtcagtattac cagatttttg ttcacatacc 66660 agtttggttt cattacctta ttggctatctggagtctagt aatgtaactg aatggaatct 66720 tgggatatta tcgctggctt tactctacatcatttaaaac ttttggcctt tttatggaca 66780 tctgagaact ttcaggtggg ttttatgaatattttttaca tgatcaagtt acggagttgc 66840 ttccagcaag agacgttttc agatgtggatgatatttgtt caatatatca agctgaattt 66900 caaagcctgt tcttaactgt cagcatgtgttttgtcaaga gtatgctacc ttatggtcta 66960 acagagggca aacatgggtg gtcaatctaaatctcatatg acccatttca cttaggttat 67020 agtgacatca gaatatgtgg gtgtgtgtatatatgtatat atatatatat atatatatat 67080 atatatatat atatgcatta gaatggatttttcagtgcta gaagttaaga aaaatgtttc 67140 caccaaatat tttagaatat tataataaagtttaagtatg ttgtacaatt tttaaaagat 67200 taaatgaatg aaagaaccct gtgttaaaaaaaattatgtt cagcctaatg tgtgtgtcag 67260 caacatttat tctatcttaa ttgacagatgattgcattgt taaactgtac atgtgtactc 67320 ttgtgttacc aaaacaactg ggtaattagaactagtgatt tagaggaaat taggtatctt 67380 ttcctgacaa tgttttcaga ataaaggatatttttcataa tattttaaga tacttgttat 67440 ctgaaagtag aattttcttt agcattggtataattcccat tctcacaaat tcttaagatc 67500 ttcataacat ttgtatttta aatgaaaattttaaatgcta tgttttatgt gtaacagatt 67560 tctaagccaa gattaattta ttgagatgacattttaatat tctctgtagg atggttagta 67620 aatttttaag acgtctttag aggttttcattgcactttgg ataaaatcta aaattcttat 67680 cattttctac aaatacttca tagtttttcttgtgcctatc actccagcct tgtttgatgc 67740 aattttctcc cctgttcatc cacataggccttcttagagt ttgtcaaggg ctctagttgt 67800 ctacctcaga gcctttatgt tgctgtgtcctagcctggaa tactcccctt ctttctctac 67860 ctttcctggc tatctccagt ttacctctcagtctcagctt aaatatcagt tcttctgata 67920 gccttccctg gttccccatc aatctaaattaagtcacttc tgctactctt tcttattatg 67980 ctttcacatt cccagtttat agttacatttatgtgtatgt gagtatatat ttaatattgg 68040 tcccttagac tgaaagctct gagaagaaaggggaaatgac tgttttgttc accattttat 68100 atgtagaacc ttttgtaatg catagcacagtgtacacatt tcataaatat ttgttaaatg 68160 aatgaataaa tgagtgaata taagattctgtgtaatctag taaaaaccca ttttcaaagt 68220 ctatgctctt ttgatccctt gtaagctgtagagttttcaa tctgtaggcc ataaatattt 68280 ctatttgtgg tcttttggaa aggaataaaaagcagtgatg attattaata tatgtaatgt 68340 atgagtaaac aatcttcatt tatatattttgctttatttt accagaatat ctttgcaagg 68400 ggtatgtgtt attgttcatg tttataccatgagaaaaata tatcagaatg gggcatactt 68460 agaaaaatga ttaatactgc ccaggaatacaaaaagagtc aaaatcttat aatatgtcac 68520 ttttacatat ttatcatagt agtttgttttgttttgtttt agatgatgca aaattgaggg 68580 gccttgcagc ccccagtaac ctttctatgaatcagactct tgaaggtcat agtggtgagt 68640 cattcaattt cttttaagtc atgtttgagtggtatttagg catttagaaa gtttctactt 68700 aatgcagaaa tgtcaacatt ttaaataatggtaatattta cagtacagat gctctttgag 68760 ttacgatggg gatatgttcc aataaacccattgtaagttg aaaatgcatt taatacacct 68820 aacctactga acatcgtagc tctgcctagcttaccttaaa tatgcccaga aaacactagc 68880 ctacagttgg gcacaatcat ctgacataaagcctatttta tactaaaatg ttgaatatct 68940 tatgtaactt attgatttct gtactaaaagtgaaaaacag aatggttgta tgggcactcg 69000 aagtacggtt tctactgaat gtgcattgcattcccatcac cgaaaagtca tttaagtagt 69060 accattgtaa gttgggagct gtttgtaatcatttctgttg agacataagt aatacaactt 69120 agagtcattc aacttttaca tgattaaacaacatcttaaa taaatctaga agaaaatagc 69180 tgtttttaat atattgaaaa tactcctgtctatggatgtt gtatatataa tcctttttag 69240 aatctgtcct atgattccag attaaaggagactgaagaga ctgggtgtag tggctaacac 69300 ctatagttca agcactttgg gaggccaaggtgggaagact gcatgaggct gggattttgg 69360 aaccaccctg ggcaacataa tgagaccctgtttctacaaa aaataaaaaa aattaaccag 69420 gcgtggtgct gtgtgcccgt agtccccagtactcaggagg ctgaggtggg aggatcgctt 69480 gagctcatga gttggaggct gcagtgagccatgattgtgc cactgcattc cagccttggt 69540 gacagtgcaa gatcctgtct caagaaaaaaaaagactaaa gactgaagat gtataacaac 69600 taaatgcact gtataatcct agattggatcttggaatgag aagaacaagt gtctataaag 69660 gacattattt ggacaattgg taaaatttgtatatggactg tgagttagat aatagtatat 69720 cagtgttaaa cttcctaatt tgtaatatgattatggaaga gaatgtcttt atttttagga 69780 aatgtacact aaagtattta gggattaaaaaattaaaagt catcaattat tcacgtaccc 69840 tagaggttta agatatgctc aagagtgttttactaataga atggaaaacg agtgagaggt 69900 gggcagaaga gagattatag gacatatctaccaaaaatga taaaagtccc ctaaacaagt 69960 acttgttatg gtgtcctttg aacatcattatttttgggag tcaggccaat tgagtaaacc 70020 atgcttgtat ttgtttctgt catcaggtatatggtagttc tggtagctag ccatgattgg 70080 tgtctttact gggatgaaat agggatagatatgaaaaata gtattatgca ttgttgtaga 70140 aaattagtga tatcaataac tgcatcttatgaataggttt tgaggaatat taaagactag 70200 ttagctgtac aggaaccata ttgattttaacatgatcttg gtgtttatat aaaaatgtga 70260 atttgaaatt aattgtaacc tttaagtttctaaattgttt tataatgttc taggttctgt 70320 tcaagttgta acatggaatg agcagtatcagaagttgact accagtgatg aaaacgggct 70380 tatcattgtg tggatgttat ataaaggtatactttactag gagagattat ttaatttttc 70440 acaagttggc atctggatca cagtatttggatgggtaaat aacatctcca aaatagaccc 70500 cttttactaa caacaggagt tcatcctgactgcacagtga taatatttct ggatttattg 70560 ctgcctactt ttcacttttt taaaactttaaccctgttat taagaaagtg aattaggtga 70620 ctgttggcca tctccaaaat gaacttagactcttagcaga agaatagctt ctacttgaga 70680 taagtctttt tttttttcca gatttccaataagaattact tctaggtagt tccagtttat 70740 aattacattc atgtgtgtgt gagtgtgtatttaatattgg ttcctcagta gactgtaagc 70800 tctgagggga aaggggaaat gactgtttgttcattttata cgtagaacct tttataatac 70860 atagcacagt gtacatgttc aataaatatttgttgaatga atgaataaat gagtggctag 70920 agttatctca gtagataaaa acacatttagtaagaagtca aagggaataa ttcacttcct 70980 gcatgggcca gttagtttcc ttttcttttatggatacaga ttttatctca actctgacta 71040 ctggttcaca gtttgtctat gagagagggtgagtaaaaga aggattaact tagataatcc 71100 acttttacca gaaaagcagc tcataatacatgttctcctg aggcctgtag ttatcatctt 71160 caaaagggaa gggtgcctta gttcatgtgtgctgttttga aaagttcatg tgtgctgttt 71220 taaatggcag ggcataccat ttaaaaagtcttcattaaag atatgctata atactttaat 71280 tttttaaaaa gtgtgtgtgt gtgtgtgtgtgtgtgtgtgt gtgtagggga tagaatgtga 71340 aagttcagtg gactggagta aactttgtctgtttccaagg taggtcttca ggtcttctct 71400 gggtctcagt ctcgattctg ttatcctgctctgggcagtt attccaaatt acatgattat 71460 ttgcttggct gcaggccaga gttgcccatgttggtatgtc aaaaagcctt tgagaaaagc 71520 aaaaacaaaa tagcctattt ttttgcttctagtttggtta tgctaaaaga caagttacat 71580 ctcattccta ccctgcatac aactagaatatttttgtcat tcggcattag agatgtcgtg 71640 tcactaatgt agtataaact gattagaattttttcaagca ggcttagggt tttacctaat 71700 atgctgtgta tttttttgcc ttcattctctttgtgatggt aactaatatg ctgcatttaa 71760 tatgcttgta attattcatt ggagagtttatgtgcatatg tataaagtta ttgttacttg 71820 gagttttttt aattggttat gatctgaaccatattgttca agtgggaaaa gttaggcttt 71880 cagaaaaatc agtagtttta gctctgaaaaagcaaatgaa gtttaatttt taattgacag 71940 aaagaaagag agagggaaag aaagaaagaagaaacaaagg atgggcattt attatgtttt 72000 ctgtgataat ctttgaagca gttatcagttaagcccaagt taatttttca ttttgttttt 72060 gttggatttg gaagttttat agtttaatgtctcaggttgt agaaaacttt gcagttattt 72120 gtcccatggc gtactttatg gaatgctttcaattcttaac tactatttaa tagatgtgaa 72180 agttttgact tgtaattttt taaatttgtgaaaacaggct cttggattga ggagatgatc 72240 aacaatcgaa ataaatcagt tgttcgcagtatgagctgga atgctgacgg acagaagatc 72300 tgcattgtat atgaagatgg ggctgtgatagttggttcag tggatggtaa tgtattgatt 72360 ggacatagaa ctaagatcaa tatctggctgacagttttct tctttagaga tttaagttag 72420 ttaccatatg ttagcattct gattgaaaaatagacataaa agggaaacta tttgatatac 72480 aatttgacga cattcaagaa ccgttctttgattttaacca tctgttttta tgtatattca 72540 aggaaatttt tttatttcct acagtggcattgaaaaaatt gaattaactt ttcagcatta 72600 ttttaaacat gtctgcctca ttatgtaatttctttttata ttggttagtc tgtacaaata 72660 tatagacttt agtttcaaaa agtctgtggtcgtttttatt ggaactgaat atgtcattgt 72720 tggttgtagt ttatagcctt aaaaacctgtatttttcaaa ttctagtaat agataaaact 72780 gggtcttctg agagtctgca cctttgttttaatttgaaac tctgtagata aataatttat 72840 tttaaaatca tttttatttc tctctctcgctcgctctgtg tgtgtgtgtg tgtgtgtgtg 72900 tctgtgtgtg tgtgtgtaac ttcatgtagtcaacaaatgt ttctgagagc ctataaagca 72960 acagctatca ttttaggaac taagggtacagcaatgacca gacctagatc aggagagatg 73020 gatgagtaac tgagaaatta cagtatcctgtgataagcgc taagatgggg aaaatacagg 73080 agcatgtaga agtgccacct aatgcagacttggagggatc aggaaaggct tcctggagga 73140 agtgatgctg atgctgaggc attagccacaaggtaggttg gtgtgtagaa tgtctctgcc 73200 ctaagaagta gtatttgagg aggtccagaggagttgcact tagctatgtt tgggccattg 73260 aaaaaagttg attatcagta gggtataatactaacagtga agagtagtat gcagtgaatc 73320 tagagagaga aacagaggcc agatcatgcagatcataagt tatattaagg tgtttggact 73380 taagtccaac agcacttggg aggccaatgaaaagttttaa gaatgggagc aatgtgataa 73440 gattgttctt tagaaacatc tctgtgctacatatgaagaa tagagaaagg aaggaaagat 73500 aatgggcagc tgtatcagga aactgctctggcaattcagg tgagagatta tggtggatgg 73560 tagccatcca ggtaatgaca agggaatggaagtaagtaga ttttatacat actcaggagg 73620 cagaatagat aggattgggg atgagaggatggaagtcccc agggcagtat ccagagtttt 73680 agcttgggca gctggatgga tggtaatgcctttcattaaa atagtgtgga gaaagattga 73740 ttttaaggga aaattcctct gttggatatgttgattttgt catgccatag gccattcaag 73800 taaatctatc cagaccatag ttagacatacatatctgcag ctcaaaaatg tctgtgctga 73860 acacaaaatt ttgggaacca ccagcatatgcatggtaatt taaaccattg aaatagttga 73920 acaaaactaa gaggcatttg tagagttaagaatcagcaac atttaggctg gggacagtgg 73980 cttatgcctg cagtcccagc actttgggaagccaaggcgg gaggattgct tgagctcagg 74040 agtttgagac cagcctggac aacatagcaagaccttgtct gtgctaaaaa taaaaaaaaa 74100 acaaaacaaa ttatctgggc gtggtggcatactcctgtag tcccaatcac ctgggaggcc 74160 aaagtgggag gattgcttga gcctgagaggttgaggctgc agtgagccat catcttacca 74220 ctacactcta gcctgggcaa aagagtgagaccctgtctca ggaaaaaaaa aaaaatcagc 74280 aacatttaag ggataggcag aagaaggatgtcgagaaata taagagtctt agagataaag 74340 gagtatatgg gctaatggca gccaagagagttcaagaagg agggagtggt aaaaaaaaaa 74400 aaaaaaaagt atcaattggc tttagcaacaggccatcagt gacgtccatg aaggcagttt 74460 ctttcttttt tttaaacaac agaaatttattttctcagag ttctagactc tagaagtcca 74520 agatcaagat gtcaacaggt ttggtttcttctttttttat ttattatttt tatagattta 74580 gagggtacaa gtgcaggttt gttatatggatatattgaat attggtgagg tctggactat 74640 tagtatataa ccataaccca aatagtgtatgttgtaggca gtttcaatgg agttgtgggg 74700 cagaagccag attgggaggg gttaagggttgagtggcagt tgaagtggag actataggtg 74760 tgaagaactc tttcatgaag tttggttgggaaaggaatgg tagaaatagt agttagaggg 74820 ggaacttggg gttaataaaa gtgtttattttatttagggt gatggtttca aagagaaata 74880 gatctgcatg tgtttaaatc cacataagaaaggccctgga gtgaagagtt taatgagaaa 74940 gagaaaatga gagtagtcga taaagggagatctttgcaaa ggtaggaagg gattagatca 75000 gagaatacat agaggaatta gccttggataggggactgac tggggaaata cctcttcttt 75060 tgtattagtg ggcaacgcag aaagtgtaagcatggactca gttatgttct gaattgttga 75120 agatgggaat gaggaatttg ttttttgggtgcttttattt tctttccaaa atagaagatg 75180 ggctcaactg ctaattggga gtgagatccagcatcagagt ttagaagaaa ggggagaaag 75240 gttgaaacag ctgctatttc agatgtcttttcaggcacat ataggtgctc agtaaatatt 75300 tattggccaa ataaattgaa tgaatgcatgaatgctgtga agaatttgtg agatctttag 75360 aaaaacaaag atttcgaaag ccgttgttggaggatcatga ccttgaacaa cttttaataa 75420 tgttttgttt taggcaatcg tatttggggaaaagacctga agggtataca gctatcccat 75480 gtaacatggt ctgcggacag taaagtcttactttttggaa tggcaaatgg ggaaatacac 75540 atttacgata atcaaggaaa ttttatggtaagtaagtgta tgcatcagtc ttgtttatac 75600 aattttatat gattatcgta catatcattcatgtttgtat atacatttat tttttatggc 75660 ggtttgtgta attaaagtct tcaattcagattttttttct ttttcccaat cgtctaaatt 75720 ttgttttaat taaataaaat tttggttttctcaaggcaaa ttagtgcaac aagttcgagt 75780 gtagctttta gttccaattt tttttggtaggtagaaagga cgaggtgtca gtgcatttaa 75840 gcagagagat aatgagtagg gcgatgtgcagtttaagttt tataggtgtc ttctggtgtc 75900 tgaagccaaa gtttacatgt agataccacagggtattttt gtagcaagct accaatggtt 75960 gtttgtaaaa agaatgacag acttaaatggtaagtgagaa aaatcaatcc ctggaaagaa 76020 ggtaaaatag aataggatca ggcagcataatctggagaaa aattctggct ttgcaaggtt 76080 tagctatcat agatgtatga ttactggtggggcagttttg ttttttagca agggttagga 76140 cattaagtaa atccactcac tggtggaagtaaaataatga acttttatgt cattttgctg 76200 ttttcaaagt acttttatat acttgttgtttgagtctcat aaaaactcag tgttttttaa 76260 tcctcatttt ttacatgtgg ggaaaatggacatttaggtg atttgtccat ggcctgtata 76320 ccagtaagtg attaagcttt aatttgccacctgattttta gactgtattt ttcagtgttc 76380 tttctacaat actctattga gtattgtctctatactgctg tgttttaaaa tgtaaaatat 76440 taaacctaac caattttatt tagataaaaatgaaactgag ttgtttggtg aatgtcactg 76500 gagctatcag cattgctgga attcattggtaccatggcac agaaggctac gtggagcctg 76560 attgcccttg ccttgctgtt tgctttgataatggaagatg ccaaataatg agacatgaga 76620 atgaccaaag taaggaattt ttctgtttaaaaaatttttt taatttctat tttttttttt 76680 ttctttttga gatggagtct cactctgtcgcccaggctgg agtgcagtgg cacgatcttg 76740 gctcacggca acttctgcct cccgggttcaagcgaatttc ctgcctcagc ctcccgagta 76800 gctgggacta cagggcgtgc cactatgcctggctaatttt ttatattttt cgtagagacg 76860 gggtttcacc ctgttagcca ggatagtctctatctcctga cctcgtgatc cacctgcctc 76920 agcctcccaa agtgctggga ttacaggcgtgagccactgt gcccggcctc aatttctaat 76980 ttttaacttt tttttttttg cgacagggtcttgctctgtt gccgaggctg gagtgcagtg 77040 gtgcgttctt ggctcactgc agccttggcctcccaggctc aaggtctcag cctcctgagg 77100 agctgggacc acaggtgtgc accaactcctccagctaatt tttgtatttt tcgtagagac 77160 agggtttcac catgttgccc aggccgatcttgaactcctg gactcaagtg atccgcccgc 77220 ctcagcctcc taaagtgttg ggattataggcgtgagccgt tgtacccagc cctgtttttt 77280 taaaaatgtg gaaagtttta aaaatgtttaaaagttgttc ttaagattat aaagttctct 77340 aggctacgta ttttcccaca gtgacctcatttttctcttt ctgactagat cccgttttga 77400 ttgacactgg catgtacgta gtaggcatccagtggaacca catgggcagc gtgttagctg 77460 tggcaggctt ccagaaggca gccatgcaggacaaagatgt gaacattgtg cagttttaca 77520 ctccgtttgg tgaggtaaat gcattcaaattgatcttctt tctttctatt taaatgtgca 77580 gtgagtaagg tagaggaaaa aataacaaaggaaacggtgt aaacttatca cattttcttt 77640 tagggacatt tcatcttcat agtataaatctttgttgtat tttatcatgg aattttataa 77700 tcttatagtt ttataattat aaaataaaaattacaatttt atacattcag aaaaccagaa 77760 gtttaacata agaaatcacc atggagtagttattttgaat attttctctg tgatctgaat 77820 atgtgtctct tttcatagcc agcttttgagtagtatttag tataatatag ttgtttcaga 77880 ttaatttttc tgatttttaa tgtgatattttaaatgtgtt tagatgttaa acatacatat 77940 ctgtctgtcc ttccttcaaa cttctaatacttttaatttg ttgcaaagta atcagttatt 78000 tgaagaaaat ttaataatta atcttaggtgatgctacatt aaaataaaag cattgttttc 78060 ataaatgtac atatgagtat tatccttctcaatagtattt ttaataaatt taattttaaa 78120 agaattttca gaagtagcat tggaaaatgtcaagtaggag aattaatata ttgtgaattt 78180 taaagaattt agaaagtgag agcttattaataactaagct taatacgttt tttgaatgtc 78240 ttattttcta acccagtact atattttgtcccttttgaat accctgttgg cttttataaa 78300 ttcgccatag actacagtga tatatatttttggctgcata aaagcctata ccagttgttt 78360 taatgttatt tttatatatt ttaataatttcttttagggc aagaaatgat gtcttctgat 78420 gcttttgaat cagtatctca aagttacccagtagaatata ctgtagaacg tagatgctct 78480 ctatacattg tcaaacgaca atatttctcgtagatgtaaa agtgaaggaa aaaaaaaagc 78540 atagttttca aggttcctaa ataattaaaaatctgcattc taattatttc tatatttata 78600 gcctccttct taattcttgt tctttattggttatttttgt tggttgccat ttacttattt 78660 atcataatgt tttatctgct tatttttgttagttcagcct tccccaggta gaaaaacatg 78720 ggttctaaag tctgtttgtt tgtcttgcaaaacaatgtgt taagtgttaa acggaaaact 78780 gtaagtgggg tttggtttag ctgtaactaaaagtatctag catagttcct caggtcatag 78840 tagtcagtat tttctgagtg agtgaatatatggggaatta tttggatctg tgttattgtt 78900 aaggtaggaa agataacctg gatggaagttatctaacaat ttaaacttaa tgatttaaac 78960 ttaagaaaac ctcagagaag aatttgcttttaaattctga gggccagggg ctcctgggtt 79020 ttgcactgtg gtcttagcca tggtaggagataacagagca tagggtttgt agtcaagttg 79080 atctggtatt gaattcattt tccacagtgtagtttgacgt agagaagtta tttagcctct 79140 gagacacagt ttagttattt tcagaaataaaggaaatacg tcatggtgct gttgtaaaga 79200 ttaaaggaga ttacaatgag gaaatgcctacacagtgcct ggcatgtaat gagggcttac 79260 taaaaggtgg ttttctttcc ttgctgctgccacatcctgg ctggtttgca gttagaagat 79320 ggtcgtattc tcctcctctt atggatgggtcaataccaag agtactgtag agggcttgct 79380 ggttcacctt taatttcata caggtagcaagacagataga tcagaccctc ctaggttaaa 79440 actttgagtt tttttcttcc attggtttccttgggaaaat ggagctcaat acatttgtct 79500 tactctatta tgtgctgcta tgacagaataccacagactg catagtatat attaacagaa 79560 acttattggc tcacagttct ggagaatgagaagtccagta ttgagctact gacatctggt 79620 gagggctttt ctgctgcata atttcatggtggaaggaaga agggcaaaga gagtgcaaca 79680 gagagacaag aggggaccta attcccctttgtataatggc agtagtccca gccatgaggg 79740 ccctcatgcc ctgatcactg cttaaaggtctcatctctta atactgtcaa aatggcaatt 79800 acatttcaat gtgagttttg gaggggacagacattcaaac tatagtaaca ttaaatttga 79860 ccctagaggg tatctgctta gaacaagggataccagtaaa gttcattgaa attttgggga 79920 ttctatcatt ctgtcaaata aaagatatagattaaaaatg ttcgattgcc ttatagccac 79980 attaaaaaag tgtttgtatt catttattcactcgttaatt tggtcataaa aagtactgtt 80040 tatttatttt aaattttaaa ttaaaaaaatttgaatgtat atttattgaa gatatatatt 80100 gactatcagt ttatcaaata atctattaatggaaagggaa agaaatgtga atatgataca 80160 gttagtactc ttaaggagct cattctgtaattctctggaa ccatagaatg ggggaagaag 80220 gcaggactcg ttgaggaagc agtatttgagctgtatcttg atggatgaat aggcgtgctc 80280 caggaggaca aggaccagta aggataggggagctgtgagg aaagaagagg gggatttcag 80340 gcagaaggga tattgtatag gccaaagctgtggaggcaga gtagttcctt cacgagcagg 80400 cacacagcag ttatggctaa atagtgggatattaagccac ataatatgat tttggacaag 80460 atgggaaaaa ggcattgtat gccatgctaaagaatgtgat ggtggggatt tactgatttt 80520 taaaagttat taaacctttt aaaaagcagggtaggttatg attagatctg tgttccagaa 80580 agataattgt gacagcagag tatagattgaattgggaggg cagaggggga gaataaggaa 80640 acagatgaaa agtaagatat aaagttaggaagctattacc acaatgcagt ccattaaaaa 80700 ggtaattgta gagagtggaa aggagtaaaagcagtaggac ttggtgactc gttggttgtg 80760 aggggatgaa agagaaagac aagactttatgattgaattt gctaataaaa tatttcattg 80820 attctaagaa atataatttt ttacaactttaaaatgagga ggcatcttat aatcaaagtt 80880 gccatatttt aattagcaat gtgttttttcctttccttag ttgtacataa aaattcatgt 80940 ttcagtctat ggcattttag atttgatgaaatttgctatt gaattctttg gatatagttt 81000 gtaaattttt cttatatgca tgtttttcattataaaaatt caagaaacgt ggctgggtgc 81060 ggtggctcac gcctgtaatc ccagcagtttgggaggccga ggcaggcaga tcatgaagtc 81120 aggagatcga gaccatcctg gctaacacagtgaaaccctg tctctactaa aaatacaaaa 81180 aattagcttg gcgtggtggc gggcgcctgtagtcccagct actcaggagg ctgaggcagg 81240 agaatggcgt gaacctggga ggcggagcttgcagtgagcc gagatcacgc cactgcactc 81300 cagcctgggt gagagtgaga aattccgtctcaaaaaaaaa aaaaaaaatt caagaaacat 81360 gattttaaag tgactagtaa cttttcacaaaataatgttc ataccacttt tgatctctgt 81420 gaagtgattg gtggtaattt gtatttcagttaattgttat actttaggtt aaagataact 81480 ttctttttca gcatctgggt actttgaaagttcctggaaa ggaaatatct gcactatctt 81540 gggaaggagg tggactgaaa attgcactagctgttgattc ctttatatat tttgcaaaca 81600 ttcgacctaa ttataaggta aatattaagagcagttgtaa aataaacagt gtttcttact 81660 aaataatttt gctatagctc aaagtatattcaaagctgtc ttgttttagg agctagcaga 81720 aaggaagcct ttttgtgtga agattttagaactagtaaac ttttctagaa tactatatat 81780 agttagtaaa attactccat gagtcttagccagaaacaga tggtacactc aaaggagtaa 81840 ttgaagagag attgatgaag ggcttattacagagatgtgg gcagggctaa agggaacagt 81900 aagggatgat aaaaagtacc acccaaagattagcgtcaat gagaagccat caccagccct 81960 gggcctgaag gggcaagaca agggagtcaggctcctggaa cccacaagag ctgtggctgt 82020 gcaagagggt ctgctctgca ggagcccagaccctgccaac cagccaggca gagagggatt 82080 tggggaataa atattccaac tcacctctccacttgctggt gctttctatt ggccaaaccc 82140 aaaggggagc cagatggcca ggtagcccaggtgacggagt tcgtagaaac tagtctccaa 82200 ggctcacggc agggtagaga agggcagagaatggattaga aggagaaaat gagaataagc 82260 aacataacat ataatcagta ttttatcttgaatttaacat ttgatccaaa agcaagctct 82320 caaatgtgag attgacctgg taaactatttgttataaaaa tcttatatat ttaattttga 82380 aaaggtaaag gaccttagag aaatggctgattctaggact agggcaggga aaatacaagg 82440 taagcctgga acagtttata gtcccagcaagtaaggaagt gctcagacaa caaaaggatg 82500 gggaaatgtc aaaaggacat tgctgaaagagcctcccaat ggccaaagct ggagcacttt 82560 gaataataaa ataaatgatg tattattttattgtgaccca aagtataaat gcccaagagt 82620 tgatgctgat atagatggat tatttgaataaataaataaa tgggagagga gaaacagatc 82680 ttccttacag aattattcca gtttacttaggtattgcctt ctccaggagg tggaacatag 82740 gcccctgttc ccctggcttg agtgtggctacacttagttt ccaaagagta gagtatggag 82800 aggggaaaaa aagtagcttt atattggtgaaacttgtcaa tcacaccttg gccaagtgat 82860 aaaggttaac atcattagga atgtcatgtggctatcacat gccccctgat atggcatgat 82920 aaagaagtgc acttcggttc tgtgctagtcttcctaaaaa cctgaagtcc aatctaagca 82980 tgaggaaaac atcagacaaa cccaaattgagggacaatct acaaaatact tgaccagtac 83040 tcctcaaaac tgttgaggtc atgaaaagcaaggaaagact gagaaattgt cacaccccag 83100 aggagcctaa ggagacatga ctaaatctaagtatcccaaa tgggattctg gaatagtgaa 83160 aggacattct ggaaaagata atgaaatctaaataaagtat gatgtttact taatagtaat 83220 agtaatatag ttaatagtga tgttgatttcctagttgtga caaatgtacc atggtaaagt 83280 aaaatgttaa caataaggga aatggatgaggagtatatgg gaactctgtg ctatctttgc 83340 aacttttctg tccatctaaa actattttaaagtaaaaaaa cttattttag aaagtaatca 83400 ggatacaaaa agctaaacat ggtttagcttagtttgtttg aatggtaaac acaatcatga 83460 taatatcaac atgatatttt atttcatttttatattttta atataattta tgtattttta 83520 tttttttatt caaaattaaa tataaacatgttgggagagt tagaagggaa agctgtaagt 83580 gtgtaagttc cataatcaat agataatgtgatacagaaat aaacaccaga acagttgcag 83640 ttaaagtagt tgtacacgac cattctagagagaggaaatt tggcaggggg ttctacattt 83700 gtttttgttt taaataagcc aggtaggattatttgaccat atattatgtg aatatgttac 83760 tctgataaaa gcaaatataa aaaatatggatgggggtaaa aaggcaaagt gactattttt 83820 gaatttcgac ttggagatcc agaattgaaactttgacagg gatcacgcta cctgttgtcc 83880 cctctccagc agttattatt gattattgaaaatttttacc attgaagctt aaatcttacc 83940 aattaaaata aggaaagtta gacaaactaaatttatattt gcagcacatt ctttgtttta 84000 taaatctcta ttatttttat aaaaggttgaatctgtcttc tatttctgaa gaatatattt 84060 tgtcagcagt gataaatatc tgcaatattttgagagtaaa aataaaataa tgtgatttgc 84120 cttcccaata ctaaataatt cttctccctttcaatctcct tccctttcct ccttcctttc 84180 ttcctagtgg ggttattgct caaacactgtagtttatgca tataccagac ctgatcgtcc 84240 agaatattgt gttgtcttct gggatacgaaaaacaatgaa aaatatgtta aatatgtgaa 84300 gggtctcatt tctattacta cctgtggagatttctgcatt ttggctacaa aagctgatga 84360 aaatcatcct caggtaggtg tttcttgatatcttaagaca tagctgggct aactgcttgg 84420 gttatagttt tctacctttt caagcatggcctctacactg gcaagtacat caaaacattg 84480 ggtctgttgg cactattggt ctagaccaattttggaaaac tctgtgactc cattgatgct 84540 gtgtccctga tttttttttt cttcaagcagctaatcttat ggcaattcat actgactggt 84600 ttaatactga agctttttgt ttctgaaatatatttaaaat accgcaatcc cttttcaaat 84660 taaaatttca tgagcaaaaa tgaacatccagatgtctact atgcataaat gctgaagaaa 84720 ttatgcagca atgaacactt gagacagtttgggtagcttt tgacattgag aaatctactt 84780 tggagttata ttgcttttaa aattaatgtatcatagcata acataaataa tataaatatg 84840 tctttttaaa aagtatacta acaacttaaatatttgcaga agtatgtaaa tatttctgaa 84900 aaggactacc aatatctttg gctgtgttatcaattatttt accccattaa aaagtataga 84960 gttttttgtt aaaggtgact ctttgaacagccatgtttgc tggtgttctc tcctgaaact 85020 tgctaaacaa tcgtaaaggt attttttaaaaagtcataag cccataggga ccagaaaggg 85080 aaaggaaatg gcaacaacat tttgtaggttagagtaggtg gattagcggt aattacctta 85140 gcagacctaa gaaagctgaa tcataagccattcagggaga aaacagaggc aactggctta 85200 gatcacggaa tcccaaagag gcacaggacttgggggtacc tggtgggaga aagaaacaaa 85260 cataaaagca atttggaagt accagagattatgcagaaaa gagaaaactt tttttaagaa 85320 gaaggagatc ccactcccct atactgagtccacaggcagc tgcccttctg ccacctacaa 85380 cgactggggt ttactctcca gagaggatcaaagagtctct gcgctgaaga gcatcaagca 85440 gagctgaggg tgggggaact ctgctgaaaacaggacaatt acgttaaaag tttgctactc 85500 aacattaccc aagtcttctt ttcctatttggttcccagaa cactggcagc caagcattga 85560 gctttgaggt gggagtttgg aaatcttctcaggagaattc gacctgttca agagaaaagc 85620 cttctaaaga tactgagatt gagagtccccagggaaatag ccctgcccat caccttacag 85680 taaagacgcc agtcagtgag gctcacttatagacaactct gttccgaaca gcttttcaat 85740 gtctgactct taaatataag cagttagacaaggattgcca gacatttgag aaagccatct 85800 aaacacaagt ggaaaagaac ttggaagaagcagagactgt gcaggaagga cacgtctgaa 85860 aaacattatt ttggggctgg gtgcgatggctcacacctgt aatcccagca ctttgagaga 85920 ccaaggcggg cggattacct gaggtcgggagttcgagatc ggcctggcta acatggtgaa 85980 accctgtctc tactaaatat acaaaattagctgggcatgg tggcgcaagc ctgtaaaccc 86040 agctactcgg gaggctgagg caggacagtcgcttgaaccc gggaggcaga ggttgcagtg 86100 agccaggatc gtgccattgc actccaacctgggcgaaagt gagactctgt ctcaaaaaaa 86160 aaaaattatt ttgaaagaac cattaatattaccttcagag agttaagaga tattacatcc 86220 ataaaataag aaaaggatac tatagaagaggaagtttttg gaaattaaaa ataggataac 86280 cataatgaaa actttggctg gaggattagaaggtaaaatg aaggaaaaca cttgtaaagt 86340 gaagcaaaag gacaaggaaa tggaaaataggggtgaaaag ataaaattag aggaccagtc 86400 cgggaagctt agcatttcaa taatatttctaaaaagagtg aacaggccac tttgggaggc 86460 cgaggtgggc agattacctg agctcaggagttcgagaccg gcctgggcaa cacggtgaaa 86520 ccctgtctct actaaaatag aaaaaattagccgggcgtgg cagcgtgtgc ctgtagtccc 86580 agctactcag gaggctgagg caggagaattgcttgaaccc aggacgcgga ggttgcggtg 86640 ggccgagatc acgccactgc actccagcctgggagacaga gtaagactct gtctccaaag 86700 aaaaaaaaaa aaaaagaaaa gaaaagaaagaaaaaagaaa aaaaaagagt gaacaggcaa 86760 aatatacaaa atacagggca gtaagtatgaaaaaaaaatt caagaaaatt ttccttgacg 86820 ttctggattg gaagtgccta gctcagtgaataaaaacatt tccacattga gacacatcat 86880 tgtagaattt cagaacattg gggagaaaacaatcttacaa gctttcaggg aggaaaaaca 86940 gatttcataa cgtcaaggat cataagtggcattctatttg tcaacagcaa tggtttggag 87000 aattgcctca aaatttgcag ggatagttttccaactagaa ttccatagct gtaagtcaac 87060 tactaattaa atatgagggt agcccagcactttgggaggc caaggtgggc agatcacctg 87120 aggttaggag tttgggacca gcctggccaacatggcaaaa ccctatctct actaaatata 87180 caaaaattag ccaggtgtgg tggcgggtgcctgtaatccc agctacttgg gaagctgagg 87240 caggagaatc gcttgaaccc tgaaggcagaggctgcagtg agccgagatg tgccactgca 87300 ctccagcctg ggcgacagag tgagactctgtctcaaagta agtaaataaa taaataaata 87360 aataaataaa taaatgggtg gaatacagacatttttagat gtgtaactca ctaaaaattt 87420 cccttccaag cttccctatc ttaggaagttactttaggaa atagtctaac aaaatgagga 87480 ataaaccaag aaagcagaag aaacagtagagtggacacca gtgaggaaga aagtaccagc 87540 tgaccatggt gcagtgtgtc cagtgagtcagatgagaggt cagaaggctc caagagaaat 87600 ttaaggggat aacaatagac tacctgacatatttgagccc ttgcagcaag atatacataa 87660 ttcagggata attggggatt tgtgttagtaataagcccat agaagactaa tcaaagagaa 87720 aaaatacaat tatcgactac agagaaagcagaatgtgctt gaaaagaaaa taaatcatga 87780 tacacttgat ggctgagttg tgaatagcatttacatagtc aaaatgatgt aatcaccctt 87840 tttaaactta aaacaccttt taattgaagtataataaaca tactgaaccc taaattctga 87900 tccgacaaaa tggtattaaa aatagagagagggatggaaa ggtgcccctg tgtggtaggg 87960 gtggggtgga aaagggaatg aaagaaagctaaagacttgt catttgtagt ggggaggcaa 88020 gagctaatgg actaaattga aataaaacaagtagcagtat aaacacgtta cttagatttg 88080 tggaggtaat agcaaaagga tcggtaaagggagcaggaaa tgggaactta gggggaactg 88140 ccatttttta gtaagaagcc ttgtggatctatttgatttt tggaactact agcatgtata 88200 actgataaaa atgaaaaata tattaaaaatacaatttgtg gtattttgtg gcagtaaata 88260 agcttatttt aagtagtgga agagtaaattatgatttttt aaaatatgat ttttattatt 88320 tggtttatat ttatgaaatt aattcaatgagttcttttta ttttttttta aatgttatgg 88380 gtaatagtag acgtatgtta tctatggggtacctgagatg ttttgagtaa ttaaacaaat 88440 tttaatcgaa tgctgaaaaa agaaaaagtctgtgtaaata gcacataaag aactgcttta 88500 tttgaaattg ttgttctagt caggaaaaacatctaagtag aatagtattc ttagtacaag 88560 atacagaaac agagactaac agtaagaaaaatgaaatgat cactttaaaa caatgtgatg 88620 taaaatgtgc attacgtttg attattttcctaatattggt cattgatgat tatttgtata 88680 taagccactt tcattaggtt gagtcatgtcttttttgagt taactgaatt tttctgaatt 88740 tctcctgtca tttatatatt gcttcaaaggaggagaacga gatggagaca tttggtgcaa 88800 cggtaacaaa tgaaattagc tcttatatagctgatcatag gataaaaatg agttttgagg 88860 ttatagctga tcagaaactg tagcaatatgaggggacttc agaaggttca tggaaaaatg 88920 gaattaacag ataaaaataa aaatagaaactttatttttc aacataagct ccatcaaggt 88980 caagacattt ttgtaagcaa tgataacagatatttagtcc atccctaaag aactgagggt 89040 cctgggaatt taaccatgtc aatgcagtcttttttacatt attaactaaa gataaatggg 89100 taccctttaa ggatttttta agattaggaaacaaaaggaa gtctggagcc aaatcaggac 89160 tgtaaagtgg atggatgcct aatgatttcccgccaaaact ctagcaaaat tgcccttgtt 89220 tgatgaatag aatgagcagg agcattgtcatggtggggag agactctggt gaagttttcc 89280 caggcatttt tctgctaaag ctttggctgactttctcaag acattctcat agtaagtgga 89340 cattattgtc ctttggccct tcagaaagtcaacaagcaaa atgccttgag catcccccaa 89400 aactgtcgcc atgacctttg ctcttggcggtctgcttttg ctttgactgg accacttcca 89460 ccacttggta accattgctt tgattgtgctttgcattcag gatcatacag gcaaagccat 89520 gtttcatctc cttttgcaat ttttgaagaaatgcctcagg ttcttgatac cacctgtttg 89580 aaatttccat ttactgctct gctcctgtgtgcagctgatc tgggtgcaac ggatttggca 89640 cccattgagt ggaaagttgg cttaactcaaatttttcagt cagaattgtg tgagctgagc 89700 cagttgagat gtctatagtg ttgactattgtttctgctgt taattgttag tcctcttcaa 89760 ttagggcaga aacaagatga gattcattttttccttgcaa attgacatgg atggtctgct 89820 gttgcaggtt tcatctttac tttgtcttgtcccttcttaa agtgagttgt ccttttgtag 89880 actgctgatt tctttagggc attgtccccataaacttttc ataaaggatt gatgatttca 89940 ccattcttcc acccaagctg caccataaatttgatgtttt ttcttgcttc aattttagca 90000 gaattaatgt ttctttgaca gggctttttttcaaactgct gccatctcct tcttagtgct 90060 tcaaactaga tcctgttcaa acatgttttaacaagttagt gcacgtttat tttgatgcaa 90120 acaattttta aatccatgct tattttttccataatacaca ttttccataa actttttgaa 90180 gaccccttgt ataaatgatg aaaagtagccagagtacttg tgtttcttgt gtcttcttaa 90240 aaaccacagt gttccattac attgagaaagcagccttggt ttagtattaa ctgatctttc 90300 tttatattta atgctgtaaa tcatctttctttttgtctgc tattttttgc acgtaacagt 90360 ttcatcagtt atttcccagt tagcaatacttgaaaaagag ataagcatga tatatattaa 90420 gcatatttcc tttattattc ataagcaggagttctgatac tttttgtttt atttgttcac 90480 agtttgtact agttctttgt aattctattggtacaccctt ggatcccaaa tacattgata 90540 ttggtaagga aatgttgata tttatttcttttccaatcag gttttatttg taacactaat 90600 ttaaaaacct aacactggta ttattaattagaatggtgtt ttctaaaacg aggatgacca 90660 tacttcctac tttgtcagat ccagtaccagtttatatctg ttaccatggt agaattctta 90720 atatcccatt ttactctcaa aagtatcctggttaagacag taaattataa ggttagtcta 90780 tctaaaatcc attataattt cacaactgctatagtaccat cattcgcttt aaatgggaaa 90840 tataattgtg taggaattgg tatgcagcgtaatgattagt atgtaataaa ctgaggatta 90900 tagaaagaat tttgctatta aaccactttcattattaata aactgcttgt aaatacctct 90960 cttccataaa aacactttgg ctgcttgtaaaagtgagttt gccctttcca gatttttcta 91020 atgaaatgag tctattaata gggacagttttaaaaattcc tgactttttg ttttcaaagg 91080 acttttgtta ctttagattt actattccctctacccaata actatgaaag taacccatta 91140 aaaattccaa atcttattct aagtagattattaaaaatca ttaacaagta ttttgttgcc 91200 atttcactgt aacttccaaa agtaatgattttcttttaat gtgtatggta gtatattgga 91260 tacagtattg taaaaattat taaatctaagaaattaaaaa cccattgcat tattcctgag 91320 aacctgagaa ctagtcattt ttttttcatattggacattt agcctaagca attaagtatt 91380 aatgcctgtt gccttttgag tgtttgcattttgctaagtg tttgttgaaa aaattgctaa 91440 gtgtttgggt gataatagca aatatgagaaataatgacct caaagacttt atactctatg 91500 gcttagggtc ttgataacct actcacttagatatattgac acatttcaac ttcatgtctt 91560 acattcattg ctacataggc atgtttatatgttttatatg tgtatatatt tatatacaca 91620 tatatataca cattagtgtt ccttgttaaaaagcttagct taaaattatt aacagactct 91680 catcacataa atatctcttg tttattagaagcattaacag tctttaagag gtaaattttg 91740 tcagtcttca gattatatgc tttttcttttaatatccaaa gttttaacat agaatcataa 91800 tggtgatagg gttaatgtca aagtaatttttatcacaagt agactgtttt tttaaaataa 91860 agctaaatgt aaatccttct tcataaaatggcaaatactt gagagcatga ctatttgtat 91920 ttttgagtct caaaactttt tttttttgtagtcttgctga ctgagatttt aacaaacagt 91980 tctttcattg gcctgcagtg aacacagccaaagactcaac atttttgttt catacatatc 92040 ctgtaaaatt tcttttttaa agtttctatgccaaatgttt atgaggacta gaggctaaaa 92100 taaaaaaagt ctgtgagtga tactcttctagaatttatta tttttgtttt gccacctact 92160 cttcctcttt ataatttttg tagaaactttaaaatgaact ttatagatta ttatagaagt 92220 gtagtttgtg ggactctgtt atactctacacatgagagtt aggtatcctt ttttggtgtt 92280 cttttcttga gcactcataa tacctttggtgtgcttccat ccaaaatacc tcttaacctg 92340 cactgtggtt actaattttc ttgtctatcttccccactag actttgaact tcttaaagat 92400 ctacgtctaa ttaacttctg tatccccatgacctaacact gccttggccc ataataaatg 92460 tttattgaat gcatggaaat tatttattttctatcatgta aataaaaaat gatgattcat 92520 ttatttagca aactatcaat gagtatatactgtttcaggt actgtactgg tcactgggaa 92580 taataagaaa ttaattatga catttcctgtgggatttctg aacaaataat taaaacctag 92640 ttaattcatt gaacagtgat ttattgcttccctactgcgt gctaggtatt gctctaggtg 92700 ccaggtaaat ggttattggt gcataaggacatcagagaca tctttctact tagagaaggt 92760 ttcatttgag ctgagttttg aaaagtaagtatttctttag gtgatatttc aaggaaagaa 92820 aacggcattt gcagaggaat gaaggcatggaaagtggcca tcaagtacca gtttgcctat 92880 gctgcagcca ttttagtggg agcacatgggcagctttcta atgatggtat ctaacacgtg 92940 cactaggttt tccatatatt ttagagaaagttaatttgat gaaaacattt cagtgaaatt 93000 tcaatttacc agcacttgaa agggctgagaaatgaataaa tgggaaagtg ggtgaacatt 93060 acagatgtag gcatttattt catgccttagcattaaaata ttacaagcag gttttattta 93120 tatgtgaatt ccaagattgt tttaaattattttacatcct tgcttggcta gtaatgactg 93180 cctattagca ttacaaagtt cagttaatgaaagcataatt attttcttaa aaactttttt 93240 tccaagctgg gcatggtggc tcactcctgtaatcccagca ctttgggagg ctgaggtggg 93300 tagatcacct gaggtcagga gttcaagaccagcctggcca aaatggcaaa accccatctc 93360 tactaaaaat acgaaaatta gccagacatggtggcatgca cctgtaatcc cagctacttg 93420 ggaggctgtg gcaggagaat cgcttgaacccaggaggcag aggttgcact gagcaaagat 93480 cgtgccacag ctctccagcc tgggtgacacagcaagacac cgtctcaaaa aaaaaaaaaa 93540 tccaataacc tttctagata tttcaaatttttttcttttc agatattgaa cagtcaaaaa 93600 gggcatggag ggccttcatc ttggctgtgttcttgtccta aaacattgta tctgttaaga 93660 ttttgttttg cttttcctag aagcttagtctagatgggat gaatcttaat aaagatctag 93720 caaaatatgt atagatagga ttatctacccatatgaattt gctgaaatta ccaccttact 93780 gattcaaaag aacatatttt tcagatatatttctattttt tgtagtaact gagaatttat 93840 atatggtttt attataaaat cctatgtaggtgataaaagt gaaaagaaca atagcaacca 93900 tgtctaacta atacttctta atccttagtgagggtcagtc tgctctagta gttaagagca 93960 gtttctggag ccagattgac caggtttgtgtccagctcta ctactttcta actgtgtgat 94020 atgggcaaat tcacatgcct cagtttctttatctgctaaa tgaggacaat aacgcctacc 94080 ttatagggaa gttgtgagtt aatatgtgttatatatttag aacactgcct gtcatatttt 94140 taaacattca ataaatactg tcttttaaaagtattattgg ccgggcgcgg tggctcacgc 94200 ctgtaatccc agcactttgg gaggccaagacgggcggatc acgaggtcag gagatcgaga 94260 ccatcctggc taacacggtg aaaccccgtctctactaaaa atacaaaaat tagccgggca 94320 tggtggcgcg cgcctgtagt cccagctacactggaggctg aggcaggaga atggcgtgaa 94380 cccgggaggc ggagcttgca gtgagtcgagaccgtgccac tgcactccag cctgggcgac 94440 agagcgaaac tccgtctcaa aaaaaaaaaaaaaaaaaagt attattaagc gtgtactcta 94500 tgccaggcac tgtgctatgc tataatttatattgtattta tatttatatt gttatctctc 94560 ttaaactttc aaacagctct gtagggttggcattattatt attttcaact tacagtaagt 94620 aaactgagag aacaaataat ttattcaaggccacagaatt aacaagtgcc tgtcatttga 94680 atccaagaag tctgactctt accttgaaccactaacattt taattttgaa gtaatcaggc 94740 taatgataaa cccttgtgag tatttaggatctatcaaaaa gaaaggaaca tgagtgtaat 94800 ttataattgt aggtaaacat tggttggaagaatgctttta tatacttagg aaaaaatgag 94860 attttattag aaagcagtta aggtgtgcaaacttgcttca aattacacta agggcataat 94920 agttaccagc attagagtat ccatggtggaatttactaaa tatttaaaaa attcttaaga 94980 tttagtgaag accaggtcag ggaaaggagagcgcagcctg acctggcagg agcgcaccct 95040 ggaggctttg ctgtcagtct ccatgcctggctttcagcaa agtgctcctt ccaagaaagg 95100 ctttgagctc cctccagagg ttggtaatagtacacatttt gaatgaaaaa atcttgacag 95160 ttacagtgaa aaaagaatta aaaaaaaaattactcagttg gactgagtta tttagcatat 95220 aaaaattatt gaatttaatc cctgagttgttaacccactc aattaacatc ataccatcag 95280 actagtttct aattctttgt tcagtttctaagtctcttgc tgcctcaaat gtcctttgtc 95340 tgcactctgg caagaactct ataaatggtacctcctagta acgttatgtc tttattccgg 95400 ccatattaat gattacattg catagtatttataatagaga aattaagttt tacactgttt 95460 tgattgttca tgaatttaga aaacaggtagtataatctgt ttgtgacgga gttttttgct 95520 tttatgtatt tcgtatgagt gtttaattttatgttgttta ttttgtgttt tgtgaccttg 95580 ggcaaattgt tctacttctt taaacctctgtttatttatc tataagatgg atttgatgag 95640 actatctact tcataaaagc atatacagtaggagctcaat taaatgttct ctaagaaatg 95700 ggcagatgaa aagaattatt tgcttaggagaatatatttc caatacatat taggaatatt 95760 aaaaattatt tgtttagaag aatatattccttaggaaggg acttgggaga tttttctttt 95820 ctaaattaga gtagaaatat aagatgtggccaggcggggt ggctcatgcc tgtaatccca 95880 gcactttggg aggctgaggc gggcagatggtgaggtcaag agatggagac aatctggcca 95940 acatggtgaa accccgtctc tactaaaaatacaaaaatta gctgggcgtg gtggtgcgtg 96000 cctgtagttt cagctactct ggaggctgaggcaggagaat tgcttgaacc cgggaggcag 96060 aggttgcaat gagctgagat cacaccactgcactccagcc tgggcgacag agtgagactc 96120 tgaaccaggt agtaattggt agtggggatgggaggaagga ctgggatttt atttattcat 96180 ttttcaatta atggttgttt gagttgtttctacttttgat cattataaat aatgttgtta 96240 tgaacatcag catatgagtt tttgtgtggacatgttttta tttttcttgg gtacatacct 96300 aggagtggaa ttgctgaggc atatagtaactctgtattta acattttgag gagttgctga 96360 actgtttacc aaagtgaaca caccatattacattcccaga tttctccata accttgctaa 96420 cacttgttat tgtctgactg tttgattctagccatcctag tgggtgtgag gtggtatctc 96480 attgtggttt tgatttgcac ttcctgatggctaatgttgt tgatcatctt tttcatgtgc 96540 ttattggcca tttgtgtatc ttcactgtagaaatgtctat ttgtatccgt tgctcacttt 96600 taaattgggc tatttgtctt tattattgagttgtaagagt tctttatatt ttctagataa 96660 aatttctttt cagatacacg atttgcaaatattttctccc attctatggg ttgtctttac 96720 tttcttgata gtgtcctttg aagcccaaatgtttttaatt ttaatgaagt ctaatttatc 96780 taatttttct tttgtcattt gtgtttttggtgtcatatct aagaagccaa ttattgcctg 96840 accaaaggtc ataaaggtgt attcccaagttttctttaag tgtttatggt tttagcattt 96900 gagtctctga tttattttga gttagtatcgttgtatgatg ctacaagctt ttgattcttt 96960 tctgatttca gagactgtgt atgggtattttgagactatg tgggggtata ttgtctataa 97020 aatgaggggt tgaactccgt gattactaaaggcctttgca gcacaagcat tctgcaattc 97080 tgtgtaactg taaattatct taatgttatttttaacagta ccattgtttg ttgcaatgac 97140 caaaacccat gtgatagcag cctcgaaagaagcattttat acctggcaat atcgtgtggc 97200 aaagaagctc acagcattgg aaattaatcagatcacacgg tctcgaaaag aagggagaga 97260 aaggtatatc ttttgataca tactttttagtagctcaacc atatcacatt gaagtcagac 97320 atagtaattg taaattgtca tgcttgataactgtagtgtc ttgattagta gaaatgtttt 97380 ttcaattaat ttgattatct ttaaaatttatgtgtttaaa tttattaaaa taaaagagaa 97440 gtagaatggt ttaataaagt ccttagagataaagagtgga aagctgtgtt tttataagga 97500 aatggctgtc ctcacttacc ctccccaccagattagagtt aatatgctat actaaaatat 97560 gattattcac ttgagatcga aggatctaactaagggttta ttcctataca ttagtggtat 97620 aggttttccc aagagggtgg ggtgattccaatatcagtga ccagaggaga ggcaggcctt 97680 tagacacaca tatggaagaa gtctttaggtggggtgaact ggggacttag cttctttttc 97740 ttgccctggt ttccaaggat tctggagttggaattacttc tagtaagtgg ggtgaaaagg 97800 gctctgccac atagtagata ggttagaagccaggatagag ggattcccca cagtaagtct 97860 gtaagtgaag aagcatttac tgtgttttgtaagttatatt gtctccatta gtgatactct 97920 gtgtttgctg ctcttctgtt gttactctaagcctgtaagt aaagcatttt gttgaatgtt 97980 ttaacctaag gattcttaat ggggacacaggcctgtgttg gtggaagtga gtggtacagt 98040 catgacagaa atgctaggtg ttttttcaaatcattatcct ccctcacatc cttgagatct 98100 taaagcagac tccccttgcc tttcctttaagactcactca tgtgctaaga tgggagatgg 98160 atcctcagta aaagtatgtt gatgggaggctgaggcagga gaatggcgtg aacccgggag 98220 gcggagcttg cagtgagccg agatcccgccactgcactcc agcctgggcg acagagcgag 98280 actccgtctc aaaaaaaaaa aaaaaaaaaaaaaaagtatg ttgagtatgc gaattcatgt 98340 ggaggcagga aaaaaagatt gaaaaatgttatatcttttg ttctcagtgg tgctggaaga 98400 ccaaatggaa agaatctttg gaaggaatctttggccagta tgtggtagtc ctcatgctgg 98460 taggcgttac cgtggtagca ataataagcagaggcctgca atcttgtggg agaaggtaga 98520 tcttaggaca aagttgaagc agaaagcctagcctctccag agtgtcttat agctgctaaa 98580 gagattatga cattagttta aatcaagaagacagcatttc cttatgtttc tgtctgggat 98640 atgtgtactg gtttgaataa ctttgagtaggttcccctta gaactcagtt tcctcatctc 98700 taaggtacag ttgatgatgg tgataacagccaacatttat tgggtacttc ttacgtgcca 98760 gacactttca taaatctaat cagtgctcattattcttgtt ttgcagagag gaaacagaca 98820 catgagctta aagtatttgc caaaggtcaccagtggaagc taaataatgt gtacacatgg 98880 acataaagtg tggaataata gacattggagactcagaagg gtgggagggt agtagggggt 98940 aagggataag aaattactta atgtgtacagtgtacattac ttgggttgtg gttgcactaa 99000 aagcccagac atcaccacta ctcaatatatccatataaca gaactgtact tttgtacccc 99060 ttaaatttat tttaaaataa taatacaggtacaggtacta gagagacagt ggatgtccaa 99120 aagtgatacc ccttcctaac acaggaaagggtaggaaagg cttctttact tcagcgcaga 99180 agttgcatta attagaacct cctaaatccacttaaatttt acttttatta aacacattga 99240 tagaagagaa attatggaga aacaacagagaaagttagtg aattagatgt tactctttac 99300 tgatgttcaa gtggagggta aggtaaaatgtctctctggg cttaatgaaa ttcatcggga 99360 atatatgatg aaatgttcag atatgtttttggtacagctt tcaggaacgt atcatttata 99420 taactagtaa acacctggta gataatttttatcttcatta agatttaact gtgttgagaa 99480 ctctctaaaa taatcagaat ttttatattaaataacagta ttaattgatg ttgtccaatc 99540 gtttattcta gagattatga aatagtataaactatactta aactttcttt gaaattctga 99600 tcttttttgc agaatttatc atgttgatgataccccttct ggatcaatgg atggtgtgct 99660 tgattatagt aaaaccattc aagtaggtgattttaatttt tccagaaaat gttaatatct 99720 gtagatttga aatattataa ttttattttgaaacctgaaa aataatttgt ttccagtgaa 99780 taggatatta ttttagcaat tctttaaaaaaatcctcttc tgtaaacttt ttcctagatc 99840 ttaaaaattc ttgaactaaa ataaagatggtagatggtga taagtagcta atatactact 99900 tagaaaagcg tattatctaa aaatgttggaaaacataact cctcctctcc tcctcacacc 99960 ttcccatctg tctatctctt tatctgtctatcagttgacc atggattggt ctgtcatccc 100020 tggcgtatta gtgagtgcat atattttgcagataatatag ttaactgttt ctttatgaaa 100080 atatgcttct gtttacaaac taatataatttgctatttca gtaaatagtt tttgaaatga 100140 ttttctcaga gtatttaaag tagtgcctattgtttataaa actgttgtct catttcttat 100200 ctgatgctca aaactctgta agtcaatcagcatgtcatct cctctcttac aggttagaaa 100260 actgttagag aagttcactg atttactcaggatcataaaa tgattaagta acaaagcaag 100320 ggtgcaagct taattttttg acagcagatctagtgatgta tctactagaa tatgcaagag 100380 ttcatctagt catctacata tccatccatttttttcaatt aagtattttt tgaatgtcta 100440 ttaagtgtga gacattggag taggtgctgggaatctactg gtgaataaga aacagtcgct 100500 ttaacaggga acttttggga aattgattcatcatcaacca gcttttctcc tgtaccctca 100560 cataaattaa aatagatgcc atcttaaaaacaacagaagt ttccattgtc aaccacgtat 100620 tttcttccag ctactacctt ctctcttcctcctttcgtag ctgagtttct tgaaagacta 100680 ttctgtattc atctttcccc tactgttcattccatctgca cttcattgca ttctggtttc 100740 cagaagactt tgaattgcca agtctgataggtctttttct gtctttatat tatttgatct 100800 acacattaca tttaatattt ttaatgactacttttttctt cttggctttc atgaaactac 100860 actcctggtt ttatctctgg cttttccttctgagtcttct ttgtcaaggg ttcatctatg 100920 catcacttat tgtgatattc cctaatgtctagccctaagc catctacctt ttttaaaaaa 100980 acctgtgaat acactcttct tagtgttcttaatgaaatcc acccctttaa tcatatgcca 101040 acaactctaa tatgtatatt tccaatgttgagttttttca atttctgaat aacatatttc 101100 ttgccatctg gatagcaaga aattgagcagctcaaaagcc ccccaagttt aatctgcctg 101160 aggctgaact cagcttccct cccaattcatttctgcatat atattctata tatcagtggt 101220 tggcttcact atctttgcaa gaatcacgagaactcatttt ctcttgcttt ccacgtctaa 101280 taaatcacca agtcctctgg attctgtttcctaaatttcc ctcaaattgg ctttcccctt 101340 tccatctctg tatcagtttg gtctaccattagtttctgcc taggtcactg caataatttc 101400 catccagcac tgcccaatgg aacttcttgtgataatggaa atactgtgtg tctttgttct 101460 ccagtgcagt agccattagc cgtatgtggctattgagtac ttaaaatatg gctagttcaa 101520 ctgaagaact gaattttaaa ttttatttcactgtcattaa ttgaagttta catttaaatg 101580 gtgacatcag gttagtggct accatatcgaacagtacaga atcgcctctc tactttcagt 101640 tttccccatc cccactatgc caatccatttattatacttc tggccactgc agcctttcta 101700 aaatataaat ctaatcatgt tacatatagttcctcccctc cccaattaca agctttctgt 101760 aagttcccat atttttcaga ataaagtataaatagccaca actactcctc cttcctggcc 101820 cacttcccag tatattcctc agataccagctgtggcaaac aacttgaaat ttcctgaatg 101880 tatcatgctg gtttttaaaa ttttttattaaaaaacattt taccttgaaa taattgtagg 101940 ctaatagaaa attgcaaaaa gagtcccttcactcagcttt cttcagtggt gtcatcttat 102000 gtaagtgtag tagagtatta aaaccaggaaactgacattg gcgcagtact gtaactagac 102060 tatgagctta ttcagttttc agttgtttttacatgtgcta ataggtgtgc gtgtgtaggt 102120 ttatctagtt ttatcaccat aagggatcttatctcttttt tcttcatttt cttttttttt 102180 tttgagacag agtcttgcac tgttgcccaggctggagtgc agtggtgtga tctcggctca 102240 ctgcagcctc cgcctcctgg gttcaagtgattctcctgcc tcagcctccc aagtagctgg 102300 gcccagtccc attcttgtcc cctggcaatcattaatatgt tctccatctc tatagttttg 102360 ttactttgag aatgttttat ggatagaatcatatagtatg tcaccttttg agattggctt 102420 tttcactaag cgtaatgccc tagagacttgtccaggttgt tgcagtgcaa tgccttctta 102480 catttcctca atttagtatg tgctggcccttctgctgtct gtttctcttt taggcctgag 102540 ctcaggtttt tgcctgtctt ggtgtaagttaggtactcct ttctctgcac tgtcataacc 102600 tctgtttctg tctcttcttt tatcatactgtcctgcaatg aggtgtttag ttcactgtct 102660 tcctcaatag acttttcagt aatctctgaagatgaagatt tcatctgagt atctctagtg 102720 cttagttcag tggttggcac atagtaggtgctcagcagta aataaatgtt ggttaagtta 102780 agaaatattg actgcttagt gaattgcatactacgttgta ccaggttagt tattcaataa 102840 atgtttgttg aatagtaatg attgattaatttattttagg gcacaaggga tccaatttgt 102900 gccataactg catcagataa gatattgattgtggtaagtt ttgcaaaaac ttattttatg 102960 aataataatt ctttaaatta taaaaatatggtaggagcca ggtgtgtgcc ccatgtctgt 103020 aatcacagtt acttgggaag ctgaagcaggaggattgctt gaggccagga aatggagacc 103080 agactgagaa catagtgaaa ccttgtctctaaaaaaaaaa attacagtaa agttttgaat 103140 atcatcttat gggcacccca agttttactgtacccttaat ataagttcag tatgagaaat 103200 aagaaagttt taaagctttc ctcgctaataattaaccatt tgcaaaaaca gctatttatg 103260 aaagcagtac agggatgacc agccgagaagacagagtaca cttacagttt ctaacactga 103320 gtggaatgag ccctgtggaa gattgcatggtggtggggca gattgcatgg tggtagtagg 103380 tattagaaag tttcctcctt tgtgtagcccgatattttat gttggagaaa gtgctttggc 103440 tcagccactt tcataagact tagaatgacgaggtaacatg tttgccagcc agtaaaatag 103500 agaagcctat ggcaaatttc agctgaatttttcaaataga atcaatggca tagtgtcttt 103560 tcttgagctc tgagatttga aaatattaattttagaaatt aatgtttact tttatacagt 103620 aatgctgctg gggttattgc atttgaattatgtaaacaaa accttaatag tatatattct 103680 agttctagtt acattcctca gccatgagctaattttatgg ggggaaaaat cttaagagca 103740 tagtatttct atagtattcc ctcctttctgataagccctc atgagactca tgctgcaaaa 103800 gcagtggttc tctgattaaa aagaagagagttaaagatca gctgtgtgga ggcttccact 103860 ttgtcccact gccatggcct gaggcagccaagggtttgaa acccacagtt ctctcatggc 103920 attatactta ctgttgatat gtcaaccttaagatttgccc tgagttgtgt acaaaattat 103980 atgctgcttt attttttaaa cttatttaaactaatattca tagtatccct gcattcagat 104040 catctttcct tttgctaaaa actagtatcagatagatgac tataggataa aaacctagac 104100 cttcagtaac agctatatca tatgatcttctgttgatttt tcatatacat atgtaattta 104160 actcaacaag tactttgctt tagcttttaatgtgccctga atattaacct agataacaaa 104220 tataaataac tttagatttt ttttccttgtgatgtaagaa tttgtgtatt gccaaagtgc 104280 taggtttctg acttctttct tgaatattacatgaaaaaaa attagaatac ttcagctgtt 104340 tgaaaaaggt ttaattaaat tagtaggtaaatatgtactt aaggacatga ttcattatca 104400 gaattgagaa tttgctgtga catcactttggttttaagac aaatgtatag acatatagag 104460 aaatagttgt agtttaatat tattatgatacacgtatgct taagtaatca aaagtaattt 104520 ccatagggtc gtgaatctgg caccattcagagatacagtc tacctaatgt tggtttgatt 104580 caaaaatatt cccttaattg tcgagcctaccagttatcct tgaattgcaa ctctaggtaa 104640 gcctgaaaac tctgctcatg tagatgttagacttagaaat gatcagttgg gattgttaga 104700 tttatatgta ataaatgtaa agatgggcgtgtctattatg aacttgtctt aaaattcagt 104760 atggttttat tttatactgc atttggtttaaatgttttta attgggcgtg tggaagtact 104820 agtataattt tttactctta aagcttaaaagcattgagac tattttatat ttcataccaa 104880 ttggttttgt ctctggatac attttcctcagaagagagca aaatagagag tagtccatat 104940 cttgtatcaa atttatttaa caaaattttaagccatttct actgtattcc aggtttctga 105000 taatctgatt ttgaagaaga atggctataggtaattattc tcatgattgc tgatagtaat 105060 tttatagttt tttccttttc tctaattgcattgtttttag ccgtcttgct atcatagaca 105120 tctcaggagt tctgactttc tttgacttggatgctcgagt aacggacagt acgggacagc 105180 aagtagttgg agagttgtta aaattggaacgaagagatgt ctgggatatg aagtgggcca 105240 aagataatcc tgatttgttt gcaatgatggagaagacaag aatgtatgtt ttcagaaact 105300 tggatcctga ggtaaaaaca agaaatgagtgttaacagtc tataaataat gagccaaata 105360 taaaaacctg gatgattccc ctttgtttctttaagaagtt tgttgaatag cagtgccaac 105420 acggctgcat gaatttctga aaataaataggaacatttct gtggatttta aaaatatttg 105480 gtcttaggtg agaaaatact tatctattaaatattctcta ctttctctga aaggtacttc 105540 ctaggcattt atttatctgt tctttttaaaaggtggagcc atggtcacta gtcttttttt 105600 cccttgtcaa acttaagaaa acaaaatcagtagccttatt taccagaatt tctgaaaaga 105660 acctaataaa aaaatatagt tttcaaatttccttgtccat tcattctttt ctgtctcttt 105720 attcacttaa ttctgtgctt actttgtcttatttatattt acatttacca tatcgttcac 105780 aaatctgtgt tctttcttaa cccagatattcttacaaata tgcaagtgca ggcagggatt 105840 aatctctctt ttgggtatga ggaagtcccctttgttctca tttttaaggt tacattttga 105900 ttttctgctt gtggctcaca tcagctaatttctatgctct ggattttcct gtcacttagt 105960 ctactctggc caactctcag ctgttgtatttaggaaatac aaaataatcc gagtggtaac 106020 ctaatcaaaa catcattaag aagataaatcttttaaatga cttttgaatt agtaatggat 106080 tttcaatttg gaattttctg tacttctgctgcactctact cttgtaaata ataaggaata 106140 agttaataaa tgctttatat cattaatttccatttgtttg catgtatagc ttccttcttg 106200 ggaagaaatt gcatgcttgg gcccctgagctgccaaaggg tttctggagt aatttattta 106260 gtggtaaact gtttgcccct ttaatatcctaaattatttt cacatttccc catttgagcc 106320 ctactcagga attttggatg taaaatgccctgatgtaaag tgctctcaaa ggctctgtta 106380 aagcttttgc aagtcttagg atctttgtgattcttcgacc ctgagaaata gttcaaagga 106440 aaggcattca gattcagggc tgctgcatttgacctgtaag catgggcttc tttcaagggt 106500 ccaagggaaa tttaatcctt gccacaaaacaattgtggga atttgacagg taatccacag 106560 gataaccttg aggatgggag tagaatactttttgaatgtt agagtctttt tctatcctta 106620 gccctgctca tactaaatca caaatcatatgcctggagaa gggagaatgt ttgttgtatc 106680 tggctgctca taatgtgtct atagcacacatggtcagtct ggtaccaagt gaaatctttg 106740 cctggtggaa agagtttctt aagtagttttaacagaacag ccttttaaaa acatactgtt 106800 ttttaatttg aatttgtttt ctggtttggcatgtagttcg atttctgtta gatttagagc 106860 tagatggatt tcttaaaacc ttcccactgtctggatttct ttataaatga tgctgttctg 106920 ttcttttcat tcacaaggac actggcttactgtggtcgca gctggtttag ctactctgct 106980 catttgcttt tctagctgct tattgatcatgtaagaacat gatctcttca aagcagggaa 107040 gatctgtttt acatactatt atatttaatagtatgttcct taaggacata atattctata 107100 ttattattta ggagataagt tattgtttgacagctaataa caatatattt ttatccattt 107160 gagtgttgtt aagaattcta gattcatacctttatctaag ctgtaaatct tcagattatt 107220 tgagagaaaa agatgccatc tgtgtgcaaagttttaattc actgataaag cacattttct 107280 ttgagaatta ggagatgaat atttaatgttgatattccct gcttgatcat tttcatctcc 107340 agtgaccctc tgaaggtact gattgcgctgttttgttctt tttcttggct cttccttctc 107400 attggcacta caatttcatt tcatgtttctttcatatctc ataatttcat caatcagact 107460 catttttgta catttgcttc tttttcaaacaaggggctcc caaacttttg aatatatgct 107520 cttatatata catatttatt tagaaattatttacatgacc taccatatta atatattatg 107580 aatcaaaata gaaattttaa aagaatgagaaaaataaatt atactggatg acataacttt 107640 atttttttaa gtcatgctaa ttgctatggttcatgctaat gctacattag ctagtatatc 107700 agcataatgc acattttaga gcaaggttttgttaacttag ttgataaaag tctcatgtgt 107760 caaatactct acttgataac tttgaccttttagctgactt ttggtcagac ctgataggat 107820 tgtcattgtt ttcacgttgt aatgtgactgatacaagaat ctgtgctagg agtaagagag 107880 gtgtcagttc tgccttttct tgtttgcttgtgcttgcatt attaatatta cttttaattt 107940 acaagttctt tgcaagaagc tttttaagccatttgtccag tttgtgagag ttagctaaaa 108000 ttaggtaata caatccctca attcacttagttaggtatat gcaaacctca gtatatcaca 108060 atatgctaaa attgtaggac aaataagggtacttcaggac actcattata tggcacatat 108120 tatgaggcct tctgacttgt gtgctggataagggtgccaa tattttgttt ggtgtagtat 108180 agattattaa agctttggtt acttttttgataaaaagaat tatcttgcat actggttagg 108240 tgagaccctg ctggtctgaa acttcatatttaaatgaagt gtgtggatag atagctaatg 108300 aattttggaa actaaagaaa attggttaattccagcagaa gttgaatttt taaaatctga 108360 attacataaa tgtatttact tatgcattaataatcagatt atttttcaaa cttttatttt 108420 ttgatctctc tttccatctt tctccctttctctttggtag tatttttttt ttcatggatc 108480 aacttttgtg agtttatctt ttgattatcttgttttgttg cactgtcttg ccaaaaataa 108540 aaaaaaaatt tctaatgata aacatgtttatacaaggatc tagggcagct actttataat 108600 atgcatatcc tattctttat ttcaacaagcctgtataaca ctaagataag agacagacat 108660 ataacagtct catgcactta catattggcaaataaatctt cttaagagta ggagatatat 108720 caggaatgca ctctggctca tgttacagaggagagactgt tcagtgtggt atgggatttc 108780 tttgatagat gggcttacat acaaatatagtgtttatcag ctttatgaag aattaacttt 108840 aatattgaat tgcatgaatt agaattttcaagctatatga ctttttatta attgagaaaa 108900 ttggactgaa tattacatgc tattagggaattaacattaa ttatgtaagg tgtgatcata 108960 gtaatgtggc catgtaagag aaaattcttatgttttaaag atatgtactg cagtatttag 109020 ggataaaata tcatgatgtc cgtaatttactgtatttact ttacaatact tcagccaaga 109080 agttaaagga agcatattta ccataatgctaattatttag gttatgggta tatgggtgtt 109140 aatgacattc tctcttttgt ttacctgaaaactctgataa taaaatttaa aagcatgatt 109200 tgttaaaaga gaggaaattt tgcataatgaaacataactt tttttttgtc tttaaaggaa 109260 cccattcaga cctctggata tatttgtaattttgaggatt tagaaattaa atctgttctt 109320 ttggatgaga tattaaaggt aattcctaaaaagttacatt tctgttgcta gcttgacagg 109380 ggattatttc tggaccaggc ggtgtagtatcctggttagc agtgtggacc caggagtcag 109440 atatatctca ggcccagccc tgctctcttattgttcagcc tttaggtggg tgccaccatc 109500 tttttgagct tcactttact ttcctgtaaaatgatgataa aaatgtataa gtctcagtta 109560 ttgcctttat aaaatacaga taataatactacttatagga ttgttattat ataagataat 109620 gtgtgaaaag caattagcac aatgctacggccaagtgaga gctaaaaaaa atgagtgcta 109680 attgttgtca ttgtttttgt agtaataacgaggtactgaa gaaatattta agagtttagc 109740 ctttttctag ttctaagata cttgagtaacttttttttag agatctatct ttccattttt 109800 tccaggtttt gtaaaataat ccttttttctctttttaaat tcaagacatg attttgaacc 109860 cctgcattgg cctacattga ccatcctttgagaatttttt ttctctaaga atatactgag 109920 agaagtttta ccttcatttt agccaaagcattaacttttt ctatgtaata tatatatagt 109980 ttttgaattg agatagtaga tttaaaatttccagaagtta gtggcaattt ggaacaggtt 110040 tgggtttgat atgtaaatca tatttgggtttgatatgtaa attatggcta ttttattgtg 110100 agccagtttc attttctata tcataaccaccatgcagcaa tgggatgggc ccccacccaa 110160 aatttggttt ggatgtaaag attgatgataccatagggac accaggaatg tatcttccat 110220 aatgaaactt ttaggggaga gaagggcaggcttcccaagc tgtcagaaat ggctggacag 110280 cacagggaaa ggagactggc ttgagggtgtttttgttgtt gttttgttgt ttagtagtta 110340 gggagtgggt ctggggtgag gcttttaatggataggggct tatgtggttt gaaactcact 110400 ggtgccccaa aggaagaagc atctgggctttctcattagc ttgccccatg tgaggcagag 110460 gggaagaggg agaggtgagg tttaaaagctgtcagcagcc aaccatcaaa aatggagtct 110520 gggctgttat tacctaggat tatgtaacctccagatttga ctatctgact cttctgtata 110580 tggatagcac ttttcttgtc cttgtttaaggtatatgact tgtaaaacta atgaacttat 110640 ttcattgccc gtttaacttg aatacagtaccaccagataa tattttttcc ttttccacac 110700 gtgtgaagaa agatatctat tgtttttttttcagcagggt ttctctctgt tgcccaggct 110760 ggagtgcagt ggtgccatca tggctcactgtagcctctgc cccccaggct caagtgatcc 110820 tcccatctca gcctcctgag tagctggggtcccaggcaca ggccgccatg cttggctaat 110880 tttttgtgtt tttttttttt tttttttgtagagatggggt ttgccatgtt ggtcaagggt 110940 ggtcttgaac tcctgggctc aagtgaaactctcattttgc cctcccaaaa tgttaggatt 111000 acaggcgtga gccaccatgc ccagccagaagatgtctttc tgcttgtctt ctttcccaag 111060 agatttaggg aaagaagaca ttatttatttatttcttttt tttttatttt gaatatgtgg 111120 ttcttccagg gccctttaaa aagacagtcataagtgggta gaatcaaact aaggtgactg 111180 aaatattttg tctgctttta tatatttaatggtttgataa agagtgagct tcttgaatcc 111240 ctaacattta gcaaggatct gatacagagttggttttcag aaagtgttgg ttaaattgat 111300 ttttaaaaat gaatacttta ctgcttttaaacaaatggca gaattatata gaattcaagc 111360 aaaacagaaa agtactgaaa caagttaaaaaaaaacccta aatcccacaa ctcagatatt 111420 ttagtcacta ccgttaaaca tcagaacatcattccttaca gctgcaagtt atatatggaa 111480 atatggattt gatagattta gatagatgaacagaagtaat tttataaaaa tgacatcatt 111540 gtacaatcta cttttttagt gttaatttaattagataacc tgaaagtgag tttaaaggaa 111600 ttctagactt ttagataata tttttccactgcaaagaata ccattttcct gggttgagaa 111660 gaaaaattat gaaaaaccag gtagttattagtttaaatta tattttaatt tttaatagta 111720 ttgtctttct actctaaaag ataacattcttacattttct tctttttctc cttttgtaga 111780 tatattattt ttacattgtc aagatttagaacatttaaat ttcgtcttgt aatcctaatt 111840 tccagttgtt tagtcttaat agttaaatgaattcaaagct attagccagg ctttttacta 111900 cattttctcc ttacgtaagt tcttaattttgattaatctt ttagttggct ggatttcatt 111960 gttagtagtg tttttcaaga aaggctcataatttccgatt gccttaagct ctagaatgtg 112020 tgaaaatgcc tgtggtgttg ttgctgtaattgatgattat attgaatttt gtgaagtttt 112080 ttagataaag aaggctctca gtggtaaaatgatttaccta aagttcatct attcattaat 112140 tcagcaacta ttaattaaca gccaataagacacagttctt ggcctcaggt agaggagttg 112200 aggagatgga taagtaattc agttgccagaaaagagttag gggtgctgtg attcagtcag 112260 gatggtacac ctgttcaaaa ttctgaagcaggagagcatt aggtcagttc tgagatgatg 112320 aatattgaat accgtaaata gtacagctaggcattgtgct gaatctttga gacatgttat 112380 ctcatttaag gtgaaaaacc aatctgagcagagggaaaca gggatgcata actgtgaggg 112440 accccttctg ccattttcct ctgtccccttttcctcatag gccgcctgat tgcctcagag 112500 gtacttactt gtgggttttg gtgttttccagataactcaa aagacccaaa cgtgaagatt 112560 tctcttcctt attttgctca caatatgatgccgttttttt tcttgttttg ttttgttctg 112620 gtttttaata taaaaaagaa acacatactctaacggcaaa tatttgaatt tttttgtttt 112680 catctttact tttaggatcc agaacatccaaacaaggatt acctaattaa ctttgagatt 112740 cggtctctgc gagatagccg agcactgattgagaaggttg gaattaaaga tgcatctcag 112800 ttcatagagg acaatccaca cccccgactttggtatttaa aaaaagcaaa ctctcccagg 112860 atgtcaggtg taggttttta aagtctcagttttgtccctc tgctccccta ctatcctcct 112920 gatgggagca gacttccatt cttgctcttctttttaagat gtagaaattc atttagattt 112980 ggaagtcctc aaaataagca gaaaattatttgaaaaatgt ttgcatatgt tactttccac 113040 catgtttctc tagaaataat agacataaatgataattgaa ttgtgattca agactagttt 113100 gtcagtaatt tgcaatatgc acttatgtttttgtttcttt taaaataaag gcgcctactg 113160 gctgaagcag ctcttcagaa actggatctatacactgcag agcaagcatt tgtgcgctgc 113220 aaagattacc aaggcattaa gtttgtgaagcgcttgggca aactactgag tgagtcaatg 113280 aaacaggctg aagttgttgg ctacttcggcaggtttgaag aggctgaaag aacgtatctc 113340 gagatggaca gaaggtaagt tatgaaggcgcaggcatccc tgagtactct atcagttcat 113400 cattgtattt tttacaaatg gaaactataaatacgttagg tgtatcatat ggttcctttt 113460 agaaattaat tttaagtttg gattacaacctagatattct acatgcctct cattcatttg 113520 tgtttcatag tctggaggca gttttcttctctgtttttac atctaataat ggctgactct 113580 gggaggtttt tttttcattt gatgggaaaaaatgatagaa caacagagac tgggatcttt 113640 tatatttaag acatagctca ttattccagttctttctttc tttgtttgtt taggccatta 113700 tttaatttta tgttgtcaag aggcataaaataaaaatcag aagtcgttag ggatcgagat 113760 tagttcttta tatttcgttt ttgatttatataaaataatg tatgaacttc ataaatattt 113820 cttcttttga attaaagaac tgtaaggcctgtctccctgg ccaggaatca aacccaggca 113880 gctgctatga aagcagataa tcttagccactgaaccacaa ggcggagagt ctgtctgaac 113940 ttcatttcag ctgttaagat tgtatctcatcttcattaat gttatttata ctcccaggag 114000 atctctaata tgacttaggc tctgaaaagtcatataaaac cacatgtgtt aacactgtgt 114060 tttgttttga aataaacctg atcacatctaagaaattgct gaaaagaaat gattgttatt 114120 taaatcatta ttttgctaaa taaattgaaagtccaggaaa gtaaaggcac aacattcttt 114180 tccctgttct ggtatcttag accatttggggtgctataac aagatacctg agactggata 114240 atttataaag aacagaaata tttttctcacagttctggag actaggaagt ctaagatcaa 114300 ggcgtcaaca agtttggtgt ctgttgagggccctgtctca ctttcaagat ggtgctttga 114360 atgttatgcc tttacatggt ggaaagtgaaaagggcaaaa ggggcaaatt ctctgaggag 114420 tcttttatga gggcattaat caattcataagggcagacat aatgacttcc cagaaacccc 114480 acctcttaat actaccacaa tggagattaagttttaacat gaattttgga ggggacacac 114540 attcaaacca taacctccag gtgctcctcatttcctctta gctagtcagg ctaagttcct 114600 aaagtgtttg gagagtcaca ttgtccactgctgtgtagcc tcagcaccta gaactgtacc 114660 taacacacag caggctcttg gtaaatatttgtcaaataga tgaatgaatg aatgaatgcc 114720 tttgtttgta gggatcttgc tattggcctccggctgaaat tgggggattg gtttagagta 114780 ctccagctcc tgaaaactgg atctggtgatgcagatgaca gtctcctgga acaagccaac 114840 aatgccattg gagactactt tgctgatcgacaaaagtggt atgtaactta cctggagctc 114900 ctcaagcatt catgcaacac ttagtagctgcctgagtact tctctaggaa gtaatcctgg 114960 ggaaatgaag atcattttct tgaattctaaaattcagata gctctagatt catcttggaa 115020 agccggagat cctgtgaagt tgtgagatgagcactataaa aatttgtagt gtataaacac 115080 aaaacttgta cttagctcca tgcagctttctgtttgagcc tacagtatat aattctaagg 115140 tgttttatgt tttatttttg aacaaagtttaataagtgtt tttatttgct ttattatata 115200 ttattttcag aaatcattgg cttttcttttcattttgatt agtatgctta tttatgtcag 115260 tgtttgggag taaagcggtg gtatttcttaagaatagtca aggaatgtta caactagaat 115320 atgctttaga tttttttccc catttatatgtttgaaagtt attctgaagc tgaagataat 115380 gaacagtaat actagtactg ggaattatgatgaaggaaca ttttggttga aagtgcttat 115440 ttatcaggag attcacatta gttttaaaagtttttccttt tttctttctg taggttgaat 115500 gctgtacaat attatgtaca aggacggaaccaggaacgct tagctgaatg ttactatatg 115560 ttagaggatt atgaagggtt agagaaccttgccatttcac ttccagaaaa ccacaagtta 115620 cttccagtag gtattgcaaa ttttagtttttggaattgtt aggttacttt ataaaatttt 115680 atgttatagt caggaaatat ctcccacaataaagaaaatt aaaaatctat aatgttacag 115740 tttcttagac atggaatgat gcttttaattataaagtaat atataaataa tatagaatgc 115800 ttgaaaatgg aggaggaaaa gtcagtcacattcttactac tttaatatga tcacagtctt 115860 gtgtagtctg ttaagattta taaagttactttttaaatgt tagctgaata catgaaatat 115920 ttacagatga aatgatatgt tgctagggatttgcttcaaa ataatctgaa attgtttgaa 115980 atttgctata acttaagttt aaaaagatagctcaactttt attatgttat attaacaata 116040 aagtatttca agttaaaaga gaaatggaaagagttctctt gctttcaaat gttcttttag 116100 agcctcttaa ttaattgttg aatactgtaaataagtttgt attttatgtg tttaatatgt 116160 gaatcactga gccgtacaat acttgtagagctagcagctc tcaactgcag agtctggatg 116220 tatatcagtt gtgagttctg cttcaaaaattttgcctatt agttaataga taagtataga 116280 attacggtaa ttcagttttc cctattgtgaatttgagcgg ctatctctgt aataatggta 116340 ttttcactca cccttatttt caatcctatgggttttcttg agtaggaaag aaactgaaat 116400 tacttagaac cctgggtata gggaagagtgtatactccgt tagtgttgtc tgttagatgt 116460 gtctgatgat gccgataaac attttgctgtagggtatcca gaatagtaat attcctgagt 116520 aaatgctaag taattataac tataatgataattaaataac taaaatagta aaaattttag 116580 tattataaat tttaaataga gttatagaagaatttttaat aatatgggaa ggtgtacata 116640 atatttaagt aggaaataca ggatacaaaattgtgtattg tgtgattgtg tacaacacaa 116700 acatatacat atatattcaa aggacaaagatgttaagaaa ttgcaccaga gtattgagag 116760 aggtttctta tgaatgataa gattatgagtaatttcactt tttcatgttt tgtaattttt 116820 tttacagata gaatgtattc ccttataatcagaaaaatat aagcattgtt ttaaaagttg 116880 gatataaaat atactttcat acttccaaacatattgtaga agggttgggg agggaaaacc 116940 tctggctaat tagatattct gaaagaaaaaaaatgatgtt ctgaaaatgg ccagtgcagc 117000 tgaacaagta ggattttttt ttgtggtggtggtggtggtg gtggtggtgg tggtggtggt 117060 ggtggtggtg gttcttcctc gctgccaccactttccaaga gagctctgtt ttgggtctag 117120 cttcctgaac tgattaatgg cagacttctctaggacattt ctttatagag gtatttgagg 117180 gctctgtgaa tgtgagattt cccttaagtctctagttaat aattagcact gagtatacca 117240 ccaattgata taaaggcttt tttcacccactggtggggga tttgttttgg agcttgacat 117300 ttctgtatac tagctactga actgagataagaaaggagtg cagtgtcttc agggcctcgt 117360 taaaaatcag atccacagcc tgagccagtgaagctctgga tttatagaag tgatgcctta 117420 gcatccattc tcatttgtac cctctgcccttgcgtctgtt tacaagaaaa atctgtaact 117480 accagtaaat ttaatactca tacgtccctacggagttttg ttaaattaaa atataagtaa 117540 tttccttggt ctgtgaaatc tgatagtattgttttgtttg tacaggaaat agcacaaatg 117600 tttgtcagag ttggaatgtg tgaacaagcagtgactgcat ttttgaaatg tagtcaacca 117660 aaggcagcag tagatacctg cgtacatctcaaccaagtag gaactgaaac tttctgtctg 117720 tgcagcttat tgtcttagga atttttaagtcaaagcaagg ttaaagtcac cctagtgatc 117780 ttttaaagat tattgccaat gtaagggttggagagcctga cgtgaatttt atgaaaggag 117840 ccaggcgggt attctcttaa ttgaggatgaagaaagttgc acttacctct taaagtgcta 117900 ccatgagggg tgacccactt ataaccagctcagtatggga gcatgaacaa accttgtgct 117960 tttagtctgg caagtttttg tctggaagtaatttcataat tttctgcatt ttgttagggt 118020 tgggttttgt ttccaaagaa ggaccagttaatggttggtc attgcctgag gaaggaattg 118080 agttacttta tcaatattta ttttggcaaatctgttttcc ttttccttta tagaaaacat 118140 acatttgata tttttaagag ctgaaattaaactaaaagta cagcgttatt atagaaacta 118200 ttgactatta atgtattctt ttaatatttttcttattttt taaaacccct tttgctttat 118260 aaaaataatt tttaaatgtc aaatctgataggacattttc acaactgaaa ttgcatattt 118320 tgttttacta taacatataa acaagtgtatctttcttgct tgaagttttg taagattaga 118380 attacaagat taatatgatt ataaactttgaaactttcat tttcacatgt atttcttaag 118440 caaatcagag tcttagagat catttaatcattttgactta tgcttatagg acttatgttg 118500 ctgtgatttt aaatgctatt tttaaaacataccagttttt aattttgaaa tagtgttttt 118560 actataaact taaagctttc ttgtgtgaattttatgatat atgtgactgt aaatacttgg 118620 gtgactgaat ggtgtaattt ttctcctagtggaacaaagc tgttgaattg gctaaaaatc 118680 atagtatgaa agaaattgga tctctgttagctaggtatgc atctcattta ctggaaaaga 118740 ataaaactct tgatgccata gaactctatcggaaagccaa ttactttttt gatgcagcta 118800 aactgatgtt taaggtaatg taaaaatcttggtgaatagt tgatttgttc cagtcacaat 118860 attttaatat atgaaagagc atattctccattttctttgg aaacaaatat ttattacagc 118920 aaaaccactc tggttttatt gcttctgtatacattatttt agtattattg ccttttacca 118980 acaaagatgt ttattaatat tagcaacttactaataccta aacaattgta tgttgaatac 119040 tggagttaac aaagagttaa agaaattactgagactgcaa agtacaaatt tctacaacag 119100 ggaattaaat tttgttgata atagagttttaagttctttt aaatgtacaa gttttaaata 119160 ttacgcttta cataatattt atttggagggaagataatta gtacagtatt aatgtataca 119220 cattttacct tctattctat tcccatttgatttaatgaaa aaggaaaaat atctcttaaa 119280 atcatttaaa taagtttcag cttctgaaggatgcaccatg ttaacactat catatctttg 119340 tgtacttgga acattatttt aaaaatctctgtattatgga aaataccagc atttcatcaa 119400 tactttcttt attctgattt agttatctaacagcacctct cattttctaa gagtgatctg 119460 aattccaact cctagggcta aatactaaggaaagaactaa atacgaatag gtactaatga 119520 aaagaagata ggttttaaaa tatttatcttaaataggtga tatttatatt ccatcttact 119580 gattttcctc cttttttagg gaatcaaatttgttagtttt ggaaatgatt gtatttgtac 119640 aaatatagat aagaagtaac ttccctcaattttgaaaaat gtttaagatt gcagatgaag 119700 aggcaaagaa aggaagtaaa cctttacgtgtcaagaagct ctatgtactg tcagccttac 119760 ttatagagca ataccatgaa cagatgaagaatgcccagcg aggaaaagtt aaaggaaaaa 119820 gttcagaggt aaagtagcac gttaaataacattacatcat tggaagtgtt taagaaaaaa 119880 aaaaaaagaa actattaaac ttcttttatggataggtcat ttaatttgca tgtctgcatt 119940 tgtttcctta aatgatatga agaaaaagaatgactgattt agaattagta ttcaaattca 120000 gaaatttcta cctagaaata ttgaaattgtttttctgtaa tactaatctg tagtctttgc 120060 tttaatgtct tatttgttaa aaaattttacttacctggga aatcttcctt tctaatcagt 120120 gagttttaaa actttcctag atatattgttgtaatgcgta aatgaatgtc tgattaactt 120180 agaatctaat tagttatatt ttaaaatttattcataccag tttaacaaac tatgaattat 120240 aataaaaaat taaatctgtt ttgggtgcagtgctcacacc agtgatctca gtgactaggg 120300 aggcttaggt gggaggatca cttgagtccatgagtttgag actgcagtga gctatgacca 120360 cgcagctgca ttccagcttg ggtgacagagtaatacctca tctcttaaaa aaaaaagtct 120420 gtaatcatag ttttgtaatg ataaaatctgctcatttaga tactttaatg actgttgtta 120480 tttgggagtt gacactgtgt gtgcgtttgtgctgacggaa agtgtgggct gactttactc 120540 gtaggccact tctgccttgg ctggtttgctggaagaagaa gttctgtcta caacagatcg 120600 tttcacagat aatgcatgga gaggggcagaggcttaccac ttctttatac ttgcacagag 120660 gcagctctat gagggatgtg tggacactgcactgaagaca ggtgggcatg tcacctaatg 120720 tgtatagtat gtctgaaaat tataaattgagtatgaaggg ctagcttctt caatttgcct 120780 ttatgtggcc ttcaataaca ttttttcctatggatttatt ttcctatgtt tgagtcttat 120840 gcattcccat ttttgaagtt ttttttaatacagagaaaac acatagttca cagaaatgtc 120900 agaaatttga gataagcaaa aagaatgtgtatagcattca tatggctagc cattcaaagt 120960 taaatacttc atagccaagt tcatattttttggagaaaaa tatatataca tatatctatt 121020 aaatatattt aatacatata tttaaatacataaaatatat ttaatacata cattcagata 121080 attttattct ttacaatatc gggagtataacatggcatac ttttatgtac tttttgaaaa 121140 ccacgtttta aaatatttcc atattttacatatacattgc tatagcattt atcttgtgtt 121200 ttataaaagt gatacttagc taataatttcaaccaaaaca gaaaaatatg aagaaaaagt 121260 aaaataatct aacaatttgt aaaataatcctataactcag aggtaatcat tgctaaggtt 121320 tgggtaaact tccttttaga tgtttctacatgcaggagca tggttttttc cctcagcagt 121380 atattatagt tctttcctca ttcttaaacagagatctata taaacattct acctaatatt 121440 ccatggtatt tgtatactgt aattgatagaagattattta catgtaatat caggatctca 121500 atttcttctt aacaataaca atattctagaatttagatat atgaactcca gagagtgtaa 121560 ttatgataat tatgttgtaa gccaaagcatatgaatgact aaatcattac tggtctttca 121620 gcttggccat ggcaagcact gtcatgacaccccacccttc ctctgtcata ttggtgacat 121680 catccatttg ctgctcacat catcaccatatgcacatcac actctaatcc taatacatca 121740 tgaatctgta ttccaatggc atgatattaaaagaaaactt ttagagaaat tcttttattt 121800 gaatatttct ggaattctat agtgatgatttttattttta attattttta taagagctct 121860 ttagagacca tatgtaataa tcgtctttcatatatattga ttttccctag catttataag 121920 ttatagttac ttgatccttt ctaagcttcagtttctcttt gtgtaaagaa aggataatag 121980 tatcttcctc agccgggtgc agtggctcatgcctgtaatc ccagcacttt gggaggctga 122040 ggcgggtgga tcacaaggtc aggagatcgagagcatcccg gctaacacgg tgaaaccctg 122100 tctttactaa aaatacaaaa aaatcagctgggtgtagtgg caggtgcctg tagtcccagc 122160 tacttgggag gctgaagcag gagaatggtgtgaacctggg aggcggagct tgcagtgagc 122220 cgagatcgca ccgctgcact ccagcctgggtgacagagcg agactccgtc tcaaaaaaaa 122280 aaaaaaatag tatcttcctc atagagttattgtaatgatt gagaagttca tgaaaagaaa 122340 actcacttat tatgctgcct ggacattgtgagccctcaat aagtggtagc tgtgatgatg 122400 atattaatag ttttttttat atagtcatgtctgtcagtct tttacttcat atttttctct 122460 taggaatcat gcttaggaat tccttcaagtttgttaaata ttatgtttat tataacatct 122520 aagttatcct gaaaccatat attaggtttattctttctta acagatttga aattctacct 122580 taatgtgccg agaccagctc agtcagggagaccctaacca agcggcgcta gaggaattaa 122640 agacacacac acagaaatat agaggtgtgaagtgggaaat caggggtctc acaaccttca 122700 gagctgagag ccccgaacag agatttacccacgtatttat taacagcaag ccagtcatta 122760 gccttgtttc tatagatatt cgattaactaaaagtatccc ttatgggaaa cgaagggatg 122820 ggctgaaata aagggatggg tctggctagttatctgcagc aggagcatgt ccttaaggca 122880 cagattgctc atgctactgg ttgtggtttaagaactcctt taagcagttt tctgccctgg 122940 gcgggccagg tgttccttgc cctcattccagtaaaccaca accttccaga gtgggtgtta 123000 tggccatcat gaacctgtca cagtgctgctgagattttgt ttatggccag ttttggggcc 123060 agtttatggc cagattttga ggggtgcctgttcccaacat gtcccccttc tttgatttgc 123120 aaatctataa aggcaaggac agctttgtcatggtgagcta cttcttgcag gagtcaggat 123180 ccacatctgc agactataga aagacaaacaacacagatta aaagcacaat catcattgaa 123240 attacagggc ttccaagtgt ttttatccattttaatggat tactagctgc taatctgtct 123300 gcagctcctt taagcactcc agttcttggcattaaggtca ggtgtccctg ggatgctttg 123360 aatatttgtt cttttaattt tgctatatccaaaaacaagt ttgtagagtg tccttctaga 123420 cgctttttta ttctttccca aattttgatcttgttaagag ctattaatag tttccacaaa 123480 tccttaatgt ttagctccta gagcgggccatatcatttga ggttgaggtg ccactatacc 123540 accatggttc tagataatag gaactcttgccgtacttctt attatatcta ccacctgaac 123600 attttgttga gaccatctga acataagtatggcatggcac acagactgag aagtgcaatt 123660 caagctaaac atccccttag gggaccaattaataatgatt ccataggaat cattgtgcag 123720 cacctctgcc tgttctgcaa tgcaatcttcctaaacaagt acgttcattt ttttctggcc 123780 aggttcaatt ttgtttacaa ataggtttttgagggcggta tgcctcaatt ataggagcag 123840 atttattatg gtaaatactg agatcagaaagctgtgtaac agcatcatag agtgattaca 123900 tctaggcatt attgccagcc aagattgataaatatgccca ataagtgtaa ttgttccctg 123960 tgtcagccct tactgaagga atactcatagcagtggtgat aacagctatc atagctacca 124020 ttaaattact cattgtgact ggttgtcccactttcctcag gttttcttcc gccatctgtg 124080 acagcttctt gatctgtccc cagttagttgactgtgttca acgggtgttg cttgcgacag 124140 ctggggtcct cctcagtgtc agtctcgaaatggctgcaac agggtggggg gggtcctcga 124200 gatcctcccg gaatctcttc cttggcatctggctcatgat aaggtttcag gtgtcttgat 124260 ggtatccaaa tcggctgttg attttggcctggagaaacac aagcataacc tctaccccaa 124320 gttattattt tacctatttc ccaactttttgttatcagat ctctccacca aactagttgt 124380 tctgcttctg tctttgcagc tggtttctgtagatgctgtt cagctgctga taacatctgg 124440 catttgggca ggctcaaaaa atttaaagttaataatgcta gattcaattg tgtatgggct 124500 gtcccataat ccctgtctct ccctcttttttgtttttgtc atcagttgtt catctgtatg 124560 aaatcataac tgagcatttt caattaactgtgtagaatga accacgtatg aagaatcaga 124620 tatcacatta ataggcaaat caaaagcagtcattacctca attacagcta caagcttcgc 124680 tttttgaact gaagtatagg gcgtctggaaaacttaacct tttgatccag aataagaagc 124740 tttaccatga ctagacccat ctgtaaaacaatgaaaacac ttagcaggct gcaggttgtt 124800 taccacagga attgtaaatg caaactgttcacagtcttgc tcagctaagg ggatagtaaa 124860 gaaacagtat tttaaatcta tgactattaaaggccaattt tttggaatta tagcaggaga 124920 aggcaatcct ggctgtaatg ctcccataggttgtataact gaattgatgg ctcttaagtc 124980 agttaaaatt ctccatttac ctgattttttcttaattacg caaactggag aattccaagg 125040 ggaaaatgtt ggagctatgt gcccattttctaattgttta gtaactaatt tctctaaagc 125100 ctccggtttc tctttactta gtggccattgttctatccaa attggcttat ctgttaacca 125160 ttttaaaggt ataggttcta gaggcttaacaatgagcacc atcaaaaatg atatcctaat 125220 ctttggcggg aactttgtct ttccacttgaagcggttctt ccaaagcttg cagatttttt 125280 tctagtccca taccagggat ataccccatttcatgcatta tatgttgact ttgagggcta 125340 tataattgtt ctggaattag aacttgtgctccccattgtt gcaataaatc tctcccccat 125400 aaatttatag gtacagaagt tataattggttgaatagtcc caggttgtcc atcgggccct 125460 tcacaatgca aaatataact actttgatatacttcagggg ctttaccaac tccaactatg 125520 ttaagttgag tgggttgaat tggccacgcagatggccagt gctgtagaga aatgattgaa 125580 atgtccgctc ctgtatctac caaacctttaaattcttttc ccctaaatag ttacttcaca 125640 ggtaggacgt ttatcagtaa tttgatttacccagtaagct gctttgcctt gtttatttgt 125700 gcttccaaat atattgtgtt ggtttaatttcacttttccc cattcccaca tatggcagaa 125760 taaggagcta tgctatgcac tctcctggcttctttctagg gaacagaagt agatataaca 125820 atttgaattt ccccattgta ttctgaatccatgactcctg tatgtattta tacccctttt 125880 aaacttaaac tagactttcc taaaagtaatcctattgtcc ccactggcaa gggtccacag 125940 accctttgcg ggggttcccc aggcagaaggctcacagttt tgtgcagcat aaatctactg 126000 cggcactact ggctgtggcg gggaacagacattgtacagg ggtgagggaa tggcctgagc 126060 tggaaatgca ccagtttgga atggggcccaggacgggccc ctcattgcat ttcccaaaat 126120 caggttccca tctttattaa acttagagtgacactgatta gcccaatgtc cttttttaca 126180 ttttggacat atttcaggct cagcagttttcttttttcag ctatctggtg gcctgacttg 126240 ctgatttttt ctacattctt ttttagtatgaccatgcttc ccacagttaa aacaccctcc 126300 aggaaatgga gtatttcctt tatccactcttagtcctgcc attgctcgtg ctagcagagt 126360 agccttatgc agattacctc cgataacatcacaggccttg ataaaatcaa ctaaatgtgc 126420 tttccctctg ataggtcgca gagcagcctggcagtcagga ttagcattgt caaaagctaa 126480 taactgcagc actatatcct gagcagccaaatctgcaatc acctttttgt tttgagatga 126540 aatctcactc tgtcgcccag cctggagtgcagtggcgcga tctcggctca ctgcaagctc 126600 cgcctcccgg gttcacgcca ttctcctgcctcagcctcct gattagctgg cactacaggc 126660 gcccgccacc acacccggcg aattttttgtatttttagta gagacggggt ttcaccatgt 126720 tagccaggat ggtctcgatc tcctgaccttgtgatccacc cccctcagcc tcccaaagtg 126780 ctgggattac aggcatgagc cactgcacccggcctgcaat caccttttta agagactcct 126840 gtaaccaagc tataaaatcc acatatggttcttttggtcc ctgttttata gcactaaagg 126900 aagcgtattg ttctccacat aaagggattttttcccaagc tctaatgcac actcctctaa 126960 gctgttctat ggcatcatcc tgcatgaccacttgtgcatc taaaccagcc cagccgccaa 127020 cccccaaaag ttggcctgta gttatattaatttgaggctg ggcctgggcg ttgcaagcag 127080 cctgaatgga agcttcatgg gcccaccaagttttaaattg taagaactga gcaggagtta 127140 gacaagcctg agtaagagca tcccagtcagtaggaatcat ctgactggaa acagcaacat 127200 tctttaacag tcccattaca aaatgagaacctggtccata ctgatttata gcttgtttaa 127260 attctttgag caatttaaaa ggaaaaggctcaaatgtagc tgtaatattt ccccgttgtt 127320 ctggggggtg tattctaaca gggaactgtcaagcctgtaa atcaccctct cgtctagctt 127380 gctgaattcc tgcctgaata gaactaagagcagtcactca aggtgctgtt cgaacagtca 127440 ctggggcaac tcttttcatc cagtgtcctctggaaaagaa agatctggag ggtcattttc 127500 ttcaaaataa taatgagggg gtgcagaaggatagggatga acctctctcc tttaccgctt 127560 tagctttagc tggcaaataa acctgctctgtaatctcttc tgttacttcg ttatactctc 127620 cttcctcctc atcatcagtg tgaaaaagttccaaggtgga acgaaccaca gcccacactt 127680 ttcccattgt taccctgatg cttccgagctccccttctta ctcactacgg ggattgcttt 127740 aagagtactc aggtgtcctc cagctagttccacgttctcc aactgttgct ccagcgaccc 127800 tttgatctgg attcaagccc ccacggtggatgccacttgc cgagaccagc tcggtcgggg 127860 agaccctaac tcagcggtgc taaaggaattaaaaacacac acacagaaat atagaggtgt 127920 gaagtgggaa atcaggggtc tcacaaccttcagagctgag agccctgaac agagatttac 127980 ccacgtattt attaacagca agccagtcattagccttgtt tctatagata tttgattaac 128040 taaaagtatc ccttatgaga aatgaaggaatgggctgaaa taaagggatg ggtctggcta 128100 gttatctgca gcaggagcat gtccttaaggcacagatcac tcatgctact ggttgtggtt 128160 taagaactcc tttaagcagt tttctgccctgggcgggcca ggtgttcctt gccctcattc 128220 cagtaaaccc acaaccttcc agcgtgggtgttatggccat catgaacctg tcacagtgct 128280 gctgagattt tgtttatggc cagttttggggctggtttat ggccagattt tgtggagcct 128340 gttcccaaca ttcatgaaac taaatttatactttttttta attatacttt aagttctggg 128400 gtacatgtgc agaatgtgca ggtttgttacataggtatac acatgcgttg gtggtttgct 128460 gcacccatca acccgtcatc tacattaggtatttctcctt atgctatccc tcccccagcc 128520 ccccaccccc tacaggcccc ggtgtgtgatctcctgcccc tgtgtccatg tgttctcatt 128580 gttcgactcc cacttatgaa tgagaacctgaggtgtttgg ttttctgttc ttgtgttagt 128640 ttgctgagaa tgatggtttc cagcttcatccacgtccctc caaaggacat gaatgcatcc 128700 ttttttatgg ctgcatagta ttccctggtgtatatgtgct gcattttctt tatccagtct 128760 atcattgatg ggcatttggg ttggctccaaatctttgcta ttgtgaacag tgctgcagta 128820 agcatacatg tgcatgtgtc tttatagtagaatgatttat aatcctttgg gtatataccc 128880 agcaatggga ttgctgggtc aaatggtatttctagttcta gatccttgag gaatcgccac 128940 actgtcttcc acaatggttg aactcatttacactcccacc aacagtgtaa aagcattcct 129000 atttctccac gtcctctcta gcatctgttgtttcctgact ttttaatgat caccattcta 129060 actggtgtga gatggtatct cattgtggttttgatttgca tttctctaat gaccagtgat 129120 gatgagcttt tcttcatgtt tgttggatgcaaaaatgtct tcttttgaaa agtttctgtt 129180 catatccttc acccactttt tgatggagttatttgttttt ttcttgtaaa tttgtttaag 129240 ttctttgtag attctggata ttagccctttgtcagatggg taggttgcaa aaattttctg 129300 taggttgcca gttcactctg ttgatagtttcttttgctgt gcagaagctc tttagtttaa 129360 tcagatccca tttttctatt ttggcttttgttgccactgc ttttggtgtt ttagttatga 129420 agtctttgcc catgcctatg tcctgaatggtattgcctag gttttcttct agggttttta 129480 tggttttagg tcttacgttt aagtctttaatccatcttga gttaattttt gtataaggtg 129540 taaggaaggg attcagtttc agctttctgcatatggctag ccagttttgc caacatcatt 129600 tattaaatag ggaatccttt ccccatttcttgtttttgtc aggtttctca aagatcagat 129660 ggttgtagat gtttggcgtt gtttctgaggcctctattct gttccattgg tctatatatc 129720 tgttttggta ccagtaccgt gccgtttttgttgctgtagc cttgtagtat agttcgtagt 129780 cagttagtgt gatgcttcca gctttgttctttttgcttag gattgtgttg gctatgcggg 129840 ctcttttttg gttccatatg aaatttaaagttctttctaa ttctgtaaag aaagtcaatg 129900 gtagcttgat tgggatagca ttgaatctataaattacttt gggcagtatg gccattttga 129960 tgatattggt tcttcctatc catgagcatggaatgttttt gcatttgttt gtgtcctctc 130020 ttattttctt gagcagtggt ttgtagttctccttgaagag gtccttcaca tcccttgtaa 130080 gttgtattcc taggtatttt attctctttgtagccattgt gaatgggagt tcactcacga 130140 tttggctctc tgtttggctg tttttggtgtataggaatgc ttgtgatttt tgcacattga 130200 ttttgtatcc tgagactttg ctgaagttacttatcagctt aaggagattt ggggctgaga 130260 tgatggagtt ttctaaatat acaatcatgtcatctgcaaa cagggacaat ttgacttcct 130320 cttttcctaa ttgaatatcc tttatttctttctcctgcct gattgtcttg gccagaactt 130380 ccaatactgt gttgaatagg agtggtgggagagggcatcc ttgtcttgtg ctggttttca 130440 aagggaatgc ttccaggtta tgcccattcagtatgatatt ggctgtgggt ttgtcataaa 130500 tggctgttat tattttgaga tatgttccatcagtgcctag tttgttgaga gtttttcgta 130560 tgaaggctgt tgaattttgt tgaaggctttttctgcatct attgagataa tcatgtggtt 130620 tatgtcactg gttttgttta tgtggtggattatatttatt gatttgcgta tgttgaacca 130680 gccttgcatc ccagggatga agccaacttgatcatggtgg ataagctttc tgatgcactg 130740 ctggatttgg tttgccagta ttttattgaggattttctca tagatgttca tcagggatat 130800 tggcctgaaa ttttttgttg ttgttgtgtctctgccacat tttggtatca ggatgatgct 130860 ggcttcataa aatgagttag ggaggattccctctttttct gttgtttgga atagtttcag 130920 aaggaatagt accagctcct ctttatacctctggtagaat ttggctgtga atccgtctgg 130980 tcctggactt tttttggttg gtgggctatgaattgctgct caatttcaga acttgttatt 131040 gatctgttca gggattcgac ttcttcctggtttagtcttg ggagggtgta tgtgtccagg 131100 aatttatcca tttcttctag attttctagtttatttgcat agaggtgttg atagtcttct 131160 ctggtggtag tctgtatttc tgtgggattggtggtgatat cccttttatc attttttatt 131220 gcatctattt gattcttctc tcttttcttctttcatagtc tagtggtcta tctattttgt 131280 tgttgtattc aaaaaaccag ttcctggattcattgatttt ttggagggtt ttttgtgtct 131340 ctatctcctt cagttctgct gtgatcttagttatttgttg tcttctgcta gcttttgaat 131400 ttgtttgctc ttgcttctct agttcttttaattgtgatgt tagggtgttg attttagatc 131460 tttcctgctt tcccatgtgg gcatttagtgctataaattt tcctctacac actgctttaa 131520 atgcattcca gaaattctgc tgcactgtgtctttgttctc attggtttca aagaacatct 131580 ttatttctgc cttcatttcg ttatttgcccagtagtcatt caggagaagg ttgttcagtt 131640 tccaggtagt tatgtggtgt tgatgagtttcttttttttt ctttttcttt ttcttttctt 131700 tttttttttt ttttttgaga tggagccttgctctgtcacc caggttggag tgcagtggtg 131760 caatctcagc tcactgcaag ctccgcctcctgggttcaca ccattctcct gcctcagcct 131820 cccaagtagc tgggactaca ggcgcccgccaccatgcctg gctaaatttt ttgtattttt 131880 agtagagatg gggttttacc atgttagccaggatggtctc aatctcctga cctcgtgacc 131940 cacccgcctt ggcctccgaa agtgctggaattacaggcgt gagccactgc gcctggcctt 132000 gatgagtttc ttaatcctga gttctaatttgattgcactg tggtctgaga gaccatttgt 132060 tatgatttct gtcttttgca tttgctgaggagtgttttac ttccaattat gtggtcagtt 132120 ttagaataac tgccatgtgg tgctgagaagaatgtatatt ctgttgattt ggggtggaga 132180 gttccgtaga tgtctattaa gttcacttggtccacagctg agttcaagtc ctgcatatcc 132240 ttgttaattt tctgtcttgt tgatctgtctaatattgaca acggggtgtt gaagtctccc 132300 actattattg tgtgggagtc taagtctctttgtaggtttc taagaactta ctttatgaat 132360 ttgggtgttc ctgtattggg tccatatatatttacgatgg ttagctcttc ttgttgcatt 132420 gatcccttta acattatgta atgcccttctttgtctcttt tgctttttgt cagttaaagt 132480 ctgttttttc agagactagg attgcaacccctgctttttt ttcttattat tgctttccat 132540 ttccttggta aatattcctc catccctttgttttgagcct atgtatgtct ttgcccgtga 132600 gatgggtctc ctgcatacag cacaccaataggtcttgact atccaatttg ccagtctgtg 132660 tcttttaatt ggggcattta gcccatttacatttaaggtt aatattgtta tgtgtgaatt 132720 tgatcctgac attatgatgc tagctggttattctgtctgt tagctgatgc agtttcttca 132780 tagcttcgat gctctttaca atttggtatgtttttgcagt ggctggtact ggttgttcct 132840 ttccatgttt agtgcttcct tcaggagctcttgtaggaga ggcctgatgg tgacaaaata 132900 tctcagcatt tccttgtctg taaaggattttatttctcct ttgcttatga agcttagttt 132960 ggttggatat gaaattctgg gttgaaaattcttttcttta agaatgttga atattggccc 133020 ccactctctt ctggctttta gggtttctgccgagagatct gctgttagtc tgatgggctt 133080 ccctttgtgg gtagcccgac ctttctctcaggttgccctt aacatttttt ccttcatttc 133140 aaccttggtg aatctgacaa tatgtgtcttggtgttgctg ttctcgagga gtatctttat 133200 ggtgttctgt gtatttcctg aatttgaatgttggcctgcc ttgctaggtt ggggattttc 133260 ctggataata tcctgaagag tgttttccaacttggttcca ttctccccat cactttcagg 133320 tacaccaatc aaacgtagat ttggccttttcacatagtcc catacttctt ggaggctttg 133380 ttcatttctt ttcactcttt tttctctaatcttgttttct tgctttattt cattgagttg 133440 atcttcaatg tctgatatcc tttcttctgcatgatcagtt tggctattga tacttgtgta 133500 tgcttcacga agttcttgtg ctgtgtttttcagctccatc aggtcattta tgttcttctc 133560 taaactggtt attctagtta gcaattcgtctaaccttttt tcaaggttct tagcttcctt 133620 gcattaggtt agaacatgct cctttaccttggaggagttt gttattaccc accttctgaa 133680 gcctgcttct gtcagttaat caaattcattctccatccag ttttgttccc ttgctggtga 133740 ggagttgtga tcctttgtag gagaagaggtgttttggttt ttggaatttt cagccttttt 133800 gctctggttt ctccccatct tcatggatttgtctaccttt ggtctttgaa gtcggtgacc 133860 ttcggatggg gtctctgagt ggacatcctttttgttggtg ttgatactat tcctttctgt 133920 ttgttagttt tccttctaac agtcaggcccctctccagca ggtctgctgg agtttgctgg 133980 aggtccgctc cagaccgttt gcctgggtatcaccagtgga ggctgcagag cagcaaaggt 134040 tgctgcctgt tcctttggaa gctttgtcccagaggggcac ctgccagatg ccagccaatg 134100 ctgtcctgta tgtggtctct gtcggcccctacatgggtcc cagggaccca ctgggaggtg 134160 tttcctagtc aggatacatg ggggtcagggacacacttga ggaggcagtc tgtcccttat 134220 cagagcttga acactgtgct ggttgatccgctgctgtctt cagagctgcc aggtagggac 134280 gtttaagtct gttggagctg cgcccataaccacccctttc cccaggtgct ttgtcccagg 134340 gaggtggaag ttttatctat aagtccctgactggggctgc tggctttttt tcagaggtgt 134400 cctgcccaga gaggaggtag tctggctgcaggggccttgc tgagctgggg tgggctccgc 134460 ccagtttcaa cttcccagcg gctttgtttacactgtgagg gtgaaaccgc ctactcaagc 134520 ctcagcaatg gtggatgcct cttcccccaccaagctcaag catcccaggt tagctcaaac 134580 tgctgtgcta accatgagac tttcaagccagtggatcttg agcttactgg gctccatggg 134640 ggtgggacct gctgagccag accccttggctccctggctt cagccccctt tccaggggag 134700 tgaatggttc tgtcttgctg gtgttccaggcgccactggg gtatgaagaa aaactcctgc 134760 agctagcttg gtgtcagctc aaacggccgcccagttttgt gctggaaacc cagggccccg 134820 gtggcgtggg cagcgaaagg aatctcctggtctgcaggtt gtgaagactg tgttaaaagt 134880 gcagtatctg ggccggagtg cacggtacagtccctaatgg cttcccttgg ctaggagagg 134940 gatttcccca tccccttgca tttcccaggtgaggtgacgc cccaccctgc tttggctcat 135000 cctccttggg ctgcacccac tgtccaactagtcccaatga gatgaacctg gtacctcagt 135060 tggaaatgcg gaaatcaccc accttctgcgtcaatcttgc tgggagctgc agaccagagc 135120 tgttcctatt cgtccatctt gccagccaaggccaactttt tttttttttt tttttttaga 135180 tggagtctcg ctctgttgcc ctggctagagtgcagtggca taatctcagc tcactgcacg 135240 ctccacgagg tcaacttttt tagatctcacatatgagtga gaacatgtga tatttgtcag 135300 aaacatctat tttagaaatc acttatgtatctgtgtttct aaagatttgt agttttcttc 135360 ttgctggctc taaaggttta tttttcagtttattcctagt tttgttgtca ttaatgaggt 135420 tgtaaatttt ttttctaatt ggttattagttatatataaa aattattttt atgtttattt 135480 tataactgac cttcatttag ttttaactaatatgtgacat aaattataca ctgtttaaaa 135540 tagctgtatc atattcaatt tatacaccatttctagactt aaaagaacaa ttagatcttt 135600 aacttgagat gcttcaccta ctaggtgaagatctctatat gcctcatttt tttttttttt 135660 gtcagtggta agacgatctg aaacaatgtatgacctgtaa attattaccc ttgaacctat 135720 catgctttgg atatgaaatt agaggatattgtttattgta actgagtgat tacattttat 135780 atacattttt tctactacac tttgttgtaaatacttgacc ccagaaccag tttataggat 135840 tctgattgta aaacacatga atatattgtttaaactggtt ttatgaagtt tttgatatac 135900 tataacaaag tatttttaag attccaaggtctagagaaga aaaagataca aattggttat 135960 aaatctaata ttaaatctta atattaaatctaatattaag tctgcttttt ttattatact 136020 ttaagttctg gggtacatgt acagaacgtgcaggcttgtt acataggtat acatgtgcca 136080 tggtggtttg ctgcacccat caaccgtcatctacattagg tatttctcct aatgctatcc 136140 ctcccctagc cccccacccc caacaggcccaagaatacag tgcattttaa agaaagattt 136200 tttcatctcc agtataattg caaataagatgttgtaactt gaattgtctc aactgacaac 136260 ttggagactc attttcactt tttcacaaaatcctattgtt tcagtgagac aaatgacaaa 136320 agcagtaata caaaagcaat aatctgtccaaagaaacaca gagaaaagaa catattattt 136380 tatactgttt tgcttatatt gttttgctttttttttaaca aaaggaaaca tggtattatc 136440 ttgcagttta tgactaacat agtataatactcatttgctt tttatgaagt gccacagaaa 136500 tttctaacat aaattgaaga aaatgtggatgttttgacac caaaattttg tgtgtgtgtg 136560 tgtatttctg agctggcaca gccatgtgcagcatttggtg gtggttcttt tcataggatg 136620 agctgttagc cagcttcaca gaaccttggatatcatgtag cctagttcct cctcttggtg 136680 ctcccatctc tcaacactgt ttgctttcagttcgttcatg caagttcaat gaaatcacct 136740 taacttagat tcttcttttg catgttcatgatatcataat aattatttta aaaaacccca 136800 attgccctgt ttttcttagc tcttcacctgaaagactatg aagacatcat ccctcctgtg 136860 gagatctact ctctgctagc actctgcgcatgcgccagca gagcctttgg gacttgttca 136920 aaagctttca ttaaacttaa atctttagagaccctcagtt cagaacagaa acagcagtat 136980 gaagaccttg ctttagaaat cttcaccaaacatacttcaa aagataacag aaaacctgaa 137040 ttggacagcc ttatggaagg gtaggctaattttaattagt ggatgccatt ctatcttatt 137100 agaagcttgg atttctagat gtacaatgtttaatgtagga attaaagcac tgaattttga 137160 agcaattcac attaacatta taccctattttatttatttt tacaagtgta ttcatgcttt 137220 attttgtcat tgtaagaaag gttttttcttgaaataactt ttttaaatga aagtatttga 137280 tgttcatctc agaagtttta ttctttaggttttttttaag tgtattaaat aaagttagac 137340 taatgagaag ttttaaagta taatgaattgtttctccccg ttttacagtg gagaagggaa 137400 actgccaaca tgcgttgcca caggaagcccaatcactgag tatcaattct ggatgtgcag 137460 tgtatgcaaa cacggtgtcc ttgctcaggaaataagccac tacagcttct gccccttatg 137520 ccatagtcca gtgggataaa ggaatgataaactgtatata actgtaaaat atatgtagca 137580 tatatacatg gctatatgct gtatatgtaataaggtttat ttctgtgagt tttgtatgaa 137640 aatcagcctc attatttata gttgaatttttgcacaaaat atatgttaat aaaataattt 137700 ttatggcaat acaactgtga aataataaggccgattttca tataaatgta tgaaaatcaa 137760 ccatgttttc tctcaccctc tctcttttgtttatcttgtt tttggttaca tgataccatg 137820 gaaatattcc aaataaattt ttttctcacagtgtcaaggt aactgaattc 137870 2 74 PRT Homo sapiens 2 Met Pro Arg Pro AlaPro Ala Arg Arg Leu Pro Gly Leu Leu Leu Leu 1 5 10 15 Leu Trp Pro LeuLeu Leu Leu Pro Ser Ala Ala Pro Asp Pro Val Ala 20 25 30 Arg Pro Gly PheArg Arg Leu Glu Thr Arg Gly Pro Gly Gly Ser Pro 35 40 45 Gly Arg Arg ProSer Pro Ala Ala Pro Asp Gly Ala Pro Ala Ser Gly 50 55 60 Thr Ser Glu ProGly Arg Ala Arg Gly Ala 65 70 3 189 PRT Homo sapiens 3 Gly Val Cys LysSer Arg Pro Leu Asp Leu Val Phe Ile Ile Asp Ser 1 5 10 15 Ser Arg SerVal Arg Pro Leu Glu Phe Thr Lys Val Lys Thr Phe Val 20 25 30 Ser Arg IleIle Asp Thr Leu Asp Ile Gly Pro Ala Asp Thr Arg Val 35 40 45 Ala Val ValAsn Tyr Ala Ser Thr Val Lys Ile Glu Phe Gln Leu Gln 50 55 60 Ala Tyr ThrAsp Lys Gln Ser Leu Lys Gln Ala Val Gly Arg Ile Thr 65 70 75 80 Pro LeuSer Thr Gly Thr Met Ser Gly Leu Ala Ile Gln Thr Ala Met 85 90 95 Asp GluAla Phe Thr Val Glu Ala Gly Ala Arg Glu Pro Ser Ser Asn 100 105 110 IlePro Lys Val Ala Ile Ile Val Thr Asp Gly Arg Pro Gln Asp Gln 115 120 125Val Asn Glu Val Ala Ala Arg Ala Gln Ala Ser Gly Ile Glu Leu Tyr 130 135140 Ala Val Gly Val Asp Arg Ala Asp Met Ala Ser Leu Lys Met Met Ala 145150 155 160 Ser Glu Pro Leu Glu Glu His Val Phe Tyr Val Glu Thr Tyr GlyVal 165 170 175 Ile Glu Lys Leu Ser Ser Arg Phe Gln Glu Thr Phe Cys 180185 4 42 PRT Homo sapiens 4 Ala Leu Asp Pro Cys Val Leu Gly Thr His GlnCys Gln His Val Cys 1 5 10 15 Ile Ser Asp Gly Glu Gly Lys His His CysGlu Cys Ser Gln Gly Tyr 20 25 30 Thr Leu Asn Ala Asp Lys Lys Thr Cys Ser35 40 5 42 PRT Homo sapiens 5 Ala Leu Asp Arg Cys Ala Leu Asn Thr HisGly Cys Glu His Ile Cys 1 5 10 15 Val Asn Asp Arg Ser Gly Ser Tyr HisCys Glu Cys Tyr Glu Gly Tyr 20 25 30 Thr Leu Asn Glu Asp Arg Lys Thr CysSer 35 40 6 42 PRT Homo sapiens 6 Ala Gln Asp Lys Cys Ala Leu Gly ThrHis Gly Cys Gln His Ile Cys 1 5 10 15 Val Asn Asp Arg Thr Gly Ser HisHis Cys Glu Cys Tyr Glu Gly Tyr 20 25 30 Thr Leu Asn Ala Asp Lys Lys ThrCys Ser 35 40 7 42 PRT Homo sapiens 7 Val Arg Asp Lys Cys Ala Leu GlySer His Gly Cys Gln His Ile Cys 1 5 10 15 Val Ser Asp Gly Ala Ala SerTyr His Cys Asp Cys Tyr Pro Gly Tyr 20 25 30 Thr Leu Asn Glu Asp Lys LysThr Cys Ser 35 40 8 37 PRT Homo sapiens 8 Ala Thr Glu Glu Ala Arg ArgLeu Val Ser Thr Glu Asp Ala Cys Gly 1 5 10 15 Cys Glu Ala Thr Leu AlaPhe Gln Asp Lys Val Ser Ser Tyr Leu Gln 20 25 30 Arg Leu Asn Thr Lys 359 18 PRT Homo sapiens 9 Leu Asp Asp Ile Leu Glu Lys Leu Lys Ile Asn GluTyr Gly Gln Ile 1 5 10 15 His Arg 10 42 PRT Homo sapiens 10 Ala Leu AspPro Cys Val Leu Gly Thr His Gln Cys Gln His Val Cys 1 5 10 15 Ile SerAsp Gly Glu Gly Lys His His Cys Glu Cys Ser Gln Gly Tyr 20 25 30 Thr LeuAsn Ala Asp Lys Lys Thr Cys Ser 35 40 11 42 PRT Homo sapiens 11 Ala LeuAsp Arg Cys Ala Leu Asn Thr His Gly Cys Glu His Ile Cys 1 5 10 15 ValAsn Asp Arg Ser Gly Ser Tyr His Cys Glu Cys Tyr Glu Gly Tyr 20 25 30 ThrLeu Asn Glu Asp Arg Lys Thr Cys Ser 35 40 12 42 PRT Homo sapiens 12 AlaGln Asp Lys Cys Ala Leu Gly Thr His Gly Cys Gln His Ile Cys 1 5 10 15Val Asn Asp Arg Thr Gly Ser His His Cys Glu Cys Tyr Glu Gly Tyr 20 25 30Thr Leu Asn Ala Asp Lys Lys Thr Cys Ser 35 40 13 228 DNA Homo sapiens 13atgccgcgcc cggcccccgc gcgccgcctc ccgggactcc tcctgctgct ctggccgctg 60ctgctgctgc cctccgccgc ccccgacccc gtggcccgcc cgggcttccg gaggctggag 120acccgaggtc ccgggggcag ccctggacgc cgcccctctc ctgcggctcc cgacggcgcg 180cccgcttccg ggaccagcga gcctggccgc gcccgcggtg caggtaca 228 14 707 DNA Homosapiens 14 aggtgtttgc aagagcagac ccttggacct ggtgtttatc attgatagttctcgtagcgt 60 acggcccctg gaattcacca aagtgaaaac ttttgtctcc cggataatcgacactctgga 120 cattgggcca gccgacacgc gggtggcagt ggtgaactat gctagcactgtgaagatcga 180 gttccaactc caggcctaca cagataagca gtccctgaag caggccgtgggtcgaatcac 240 acccttgtca acaggcacca tgtcaggcct agccatccag acagcaatggacgaagcctt 300 cacagtggag gcaggggctc gagagccctc ttctaacatc cctaaggtggccatcattgt 360 tacagatggg aggccccagg accaggtgaa tgaggtggcg gctcgggcccaagcatctgg 420 tattgagctc tatgctgtgg gcgtggaccg ggcagacatg gcgtccctcaagatgatggc 480 cagtgagccc ctagaggagc atgttttcta cgtggagacc tatggggtcattgagaaact 540 ttcctctaga ttccaggaaa ccttctgtgg taagttggtc agtctttgcttccaactaga 600 aaggatgtta tattcagaca ttgtgtaccc agagaaaaga gtattattctcttttttttg 660 gtggaaattt aatgaaaaac ttaaatatat ccaatgcgtg tgtgtgt 70715 326 DNA Homo sapiens 15 catgggagac ggtatctgag atgtttccta agtgtcagtgctgactgttg taaacttcct 60 ctcacagcgc tggacccctg tgtgcttgga acacaccagtgccagcacgt ctgcatcagt 120 gatggggaag gcaagcacca ctgtgagtgt agccaaggatacaccttgaa tgccgacaag 180 aaaacgtgtt caggtgaggc ctgtgtaggg ggccgtgactgaatcaaata cttgggaacc 240 tatgctccag gttttggact ggccttgtgc tttgactctctgaccccttt ttacctaact 300 gccttttggc tggtttactg gtgttc 326 16 261 DNAHomo sapiens 16 ccctcccttg tttcattagc tctgtgtgtg tgtgtgtgtg tgtgtgtgtgtgtataactg 60 taagtggcgt ggcccacaat tatttctgcc atgctctgca tgcatttacatttgtcccaa 120 tcatcttttg ataagctctt gataggtgtg ctcttaacac ccacggatgtgagcacatct 180 gtgtgaatga cagaagtggc tcttatcatt gtgagtgcta tgaaggttataccttgaatg 240 aagacaggaa aacttgttca g 261 17 130 DNA Homo sapiens 17agctcaagat aaatgtgctt tgggtaccca tgggtgtcag cacatttgtg tgaatgacag 60aacagggtcc catcattgtg aatgctatga gggctacact ctgaatgcag ataaaaaaac 120atgttcaggt 130 18 130 DNA Homo sapiens 18 agtccgtgac aagtgtgccctaggctctca tggttgccag cacatttgtg tgagtgatgg 60 ggccgcatcc taccactgtgattgctatcc tggctacacc ttaaatgagg acaagaaaac 120 atgttcaggt 130 19 199DNA Homo sapiens 19 agccactgag gaagcacgaa gacttgtttc cactgaagatgcttgtggat gtgaagctac 60 actggcattc caggacaagg tcagctcgta tcttcaaagactgaacacta aacatatcct 120 ttctggaagg ttttctcctt ctgcctggag ccaaccaaggtactacgatc gagccaaaca 180 ctttaagaca ttccacagg 199 20 327 DNA Homosapiens 20 gtttgaaagt gtgtttatat tttctaatgt gtaaaaataa atggtgttggccttaacaac 60 ttgccacttg atgacatttt ggagaagttg aaaataaatg aatatggacaaatacatcgt 120 taaattgctc caatttctca cctgaaaatg tggacagctt ggtgtacttaatactcatgc 180 attcttttgc acacctgtta ttgccaatgt tcctgctaat aatttgccattatctgtatt 240 aatgcttgaa tattactgga taaattgtat gaagatcttc tgcagaatcagcatgattct 300 tccaaggaaa tacatatgca gatactt 327 21 22 DNA ArtificialSequence primer that hybridizes to the human MATN3 gene 21 ttaaacaagagcccacagca aa 22 22 22 DNA Artificial Sequence primer that hybridizes tothe human MATN3 gene 22 ttaaacaaga gcccacagca aa 22 23 21 DNA ArtificialSequence primer that hybridizes to the human MATN3 gene 23 gcgaggctcggatttaaagg t 21 24 18 DNA Artificial Sequence primer that hybridizes tothe human MATN3 gene 24 gacgcgtgag catcgaac 18 25 22 DNA ArtificialSequence primer that hybridizes to the human MATN3 gene 25 caggaatcaattgctgacca aa 22 26 22 DNA Artificial Sequence primer that hybridizes tothe human MATN3 gene 26 cgtaggatac cccaggatca ag 22 27 22 DNA ArtificialSequence primer that hybridizes to the human MATN3 gene 27 ccatttcaaatctgccactt ca 22 28 23 DNA Artificial Sequence primer that hybridizes tothe human MATN3 gene 28 cccttctgca tctttctgga taa 23 29 22 DNAArtificial Sequence primer that hybridizes to the human MATN3 gene 29tagcagcaat ttgattccca ag 22 30 20 DNA Artificial Sequence primer thathybridizes to the human MATN3 gene 30 ttgctaagct tcgctatgga 20 31 20 DNAArtificial Sequence primer that hybridizes to the human MATN3 gene 31gagggaggca gatgaaagaa 20 32 20 DNA Artificial Sequence primer thathybridizes to the human MATN3 gene 32 aggccttctc ccattgagtt 20 33 23 DNAArtificial Sequence primer that hybridizes to the human MATN3 gene 33tccagaattt tacttcattc ctg 23 34 26 DNA Artificial Sequence primer thathybridizes to the human MATN3 gene 34 tgaagagatt tattctgaga caaaca 26 3518 DNA Artificial Sequence primer that hybridizes to the human MATN3gene 35 atgggttgat gggtgcag 18 36 20 DNA Artificial Sequence primer thathybridizes to the human MATN3 gene 36 tctgggatac atgtgcagaa 20 37 21 DNAArtificial Sequence primer that hybridizes to the human MATN3 gene 37aacaactcca gcagccataa a 21 38 23 DNA Artificial Sequence primer thathybridizes to the human MATN3 gene 38 gcttttagag actgggtctc att 23 39 20DNA Artificial Sequence primer that hybridizes to the human MATN3 gene39 gtttttagcc tgctgcttgg 20 40 20 DNA Artificial Sequence primer thathybridizes to the human MATN3 gene 40 tgttggtacc ttgcgattcc 20 41 18 DNAArtificial Sequence primer that hybridizes to the human MATN3 gene 41ctgcaaccac cacctcct 18 42 18 DNA Artificial Sequence primer thathybridizes to the human MATN3 gene 42 ggcgcacacc tgtagtcc 18 43 42 PRTHomo sapiens 43 Val Arg Asp Lys Cys Ala Leu Gly Ser His Gly Cys Gln HisIle Cys 1 5 10 15 Val Ser Asp Gly Ala Ala Ser Tyr His Cys Asp Cys TyrPro Gly Tyr 20 25 30 Thr Leu Asn Glu Asp Lys Lys Thr Cys Ser 35 40 44 42PRT Mus musculus 44 Ala Leu Asp Gln Cys Met Leu Gly Thr His Gln Cys GlnHis Val Cys 1 5 10 15 Val Ser Asp Gly Asp Gly Lys His His Cys Glu CysSer Gln Gly Tyr 20 25 30 Thr Leu Asn Ala Asp Gly Lys Thr Cys Ser 35 4045 42 PRT Mus musculus 45 Ala Ile Asp Lys Cys Ala Leu Ser Thr His GlyCys Glu Gln Ile Cys 1 5 10 15 Ile Asn Asp Arg Asn Gly Ser Tyr His CysGlu Cys Tyr Gly Gly Tyr 20 25 30 Ala Leu Asn Ala Asp Arg Arg Thr Cys Ala35 40 46 42 PRT Mus musculus 46 Ala Leu Asp Lys Cys Ala Ser Gly Thr HisGly Cys Gln His Ile Cys 1 5 10 15 Val Asn Asp Gly Ala Gly Ser His HisCys Glu Cys Phe Glu Gly Tyr 20 25 30 Thr Leu Asn Ala Asp Lys Lys Thr CysSer 35 40 47 42 PRT Mus musculus 47 Val Arg Asn Lys Cys Ala Leu Gly ThrHis Gly Cys Gln His Ile Cys 1 5 10 15 Val Ser Asp Gly Ala Val Ala TyrHis Cys Asp Cys Phe Pro Gly Tyr 20 25 30 Thr Leu Asn Asp Asp Lys Lys ThrCys Ser 35 40 48 41 PRT Gallus gallus 48 Ala Ala Asn Thr Cys Ala Leu GlyThr His Asp Cys Glu Gln Val Cys 1 5 10 15 Val Ser Asn Asp Gly Ser TyrLeu Cys Asp Cys Tyr Glu Gly Tyr Thr 20 25 30 Leu Asn Pro Asp Lys Arg ThrCys Ser 35 40 49 41 PRT Gallus gallus 49 Ala Val Asp Val Cys Ala Pro GlyArg His Glu Cys Asp Gln Ile Cys 1 5 10 15 Val Ser Asn Asn Gly Ser TyrVal Cys Glu Cys Phe Glu Gly Tyr Thr 20 25 30 Leu Asn Pro Asp Lys Lys ThrCys Ser 35 40 50 41 PRT Gallus gallus 50 Ala Met Asp Val Cys Ala Pro GlyArg His Asp Cys Ala Gln Val Cys 1 5 10 15 Arg Arg Asn Gly Gly Ser TyrSer Cys Asp Cys Phe Glu Gly Phe Thr 20 25 30 Leu Asn Pro Asp Lys Lys ThrCys Ser 35 40 51 41 PRT Gallus gallus 51 Ala Val Asp Val Cys Ala Pro GlyArg His Asp Cys Glu Gln Val Cys 1 5 10 15 Val Arg Asp Asp Leu Phe TyrThr Cys Asp Cys Tyr Gln Gly Tyr Val 20 25 30 Leu Asn Pro Asp Lys Lys ThrCys Ser 35 40 52 55 DNA Homo sapiens 52 aggctgcagc cgcggatggc tgccccccctgccccggtgt tactgggagg acgga 55 53 54 DNA Homo sapiens 53 aggctgcagccgcggatggc tgcccccccg ccccggtgtt actgggagga cgga 54 54 50 DNA Homosapiens 54 cgggccggcc tggcgctcga ggccccgggg cggggcgggg cccgaggccg 50 5560 DNA Homo sapiens 55 cgggccggcc tggcgctcga ggcccggggg cggggccggggcggggcggg gcccgaggcc 60 56 50 DNA Homo sapiens 56 actttttttt tgagatgatgtcttgctctt attgcccagg ctggagtgca 50 57 51 DNA Homo sapiens 57 actttttttttgagatgatg tcttggctct tattgcccag gctggagtgc a 51 58 50 DNA Homo sapiens58 aaaccttctg tggtaagttg gtcagtcttt gcttccaact agaaaggatg 50 59 48 DNAHomo sapiens 59 aaaccttctg tggtaagttg gtcagtttgc ttccaactag aaaggatg 4860 54 DNA Homo sapiens 60 gtgtacccag agaaaagagt attattcttt tttggtggaaatttaatgaa aaac 54 61 50 DNA Homo sapiens 61 gtgtacccag agaaaagagtattatttttg gtggaaattt aatgaaaaac 50 62 53 DNA Homo sapiens 62 catgggagacggtatctgag atgtttccta agtgtcagtg ctgactgttg taa 53 63 51 DNA Homosapiens 63 catgggagac ggtatctgag atgttataag tgtcagtgct gactgttgta a 5164 64 DNA Homo sapiens 64 aaagaatact agtgggaaag ttagttatat atatatatatttttttttct tctctttttt 60 gaga 64 65 66 DNA Homo sapiens 65 aaagaatactagtgggaaag ttagttatat atatatatat attttttttt cttctctttt 60 ttgaga 66 6672 DNA Homo sapiens 66 cctgtatcat ttccccatct gtgcttgtgt gtgtgtgtgtgtgtgtgcat gtgtctcctg 60 gtcagtcttt tc 72 67 74 DNA Homo sapiens 67cctgtatcat ttccccatct gtgcttgtgt gtgtgtgtgt gtgtgtgtgc atgtgtctcc 60tggtcagtct tttc 74 68 50 DNA Homo sapiens 68 tataattctt ttggctttgaaacttgcaac tttgagaaca aaacagtcct 50 69 52 DNA Homo sapiens 69 tataattcttttggctttga aacttttgca actttgagaa caaaacagtc ct 52 70 20 DNA ArtificialSequence primer that hybridizes to the human MATN3 gene 70 agaatgagggatgccaactt 20 71 18 DNA Artificial Sequence primer that hybridizes tothe human MATN3 gene 71 gccaaggtgg gaggatca 18 72 22 DNA ArtificialSequence primer that hybridizes to the human MATN3 gene 72 tgagggatgccaacttaatc tt 22 73 20 DNA Artificial Sequence primer that hybridizes tothe human MATN3 gene 73 gacatgctca tgaagctgga 20 74 19 DNA ArtificialSequence primer that hybridizes to the human MATN3 gene 74 cctggtaatcccaacactg 19 75 20 DNA Artificial Sequence primer that hybridizes to thehuman MATN3 gene 75 cagctcactg caagctctgt 20 76 20 DNA ArtificialSequence primer that hybridizes to the human MATN3 gene 76 ccaagcaatgatcagaccac 20 77 24 DNA Artificial Sequence primer that hybridizes tothe human MATN3 gene 77 tcagacattg tgtacccaga gaaa 24 78 22 DNAArtificial Sequence primer that hybridizes to the human MATN3 gene 78ctggtccctg aatcctgttc tt 22 79 22 DNA Artificial Sequence primer thathybridizes to the human MATN3 gene 79 ctattccaat gtcaccgtca gc 22 80 22DNA Artificial Sequence primer that hybridizes to the human MATN3 gene80 cccaccccca gaatgttact ta 22 81 24 DNA Artificial Sequence primer thathybridizes to the human MATN3 gene 81 tcagcatgca tttattgatc tctg 24 8222 DNA Artificial Sequence primer that hybridizes to the human MATN3gene 82 ccagaaggaa aagattgcca ct 22 83 22 DNA Artificial Sequence primerthat hybridizes to the human MATN3 gene 83 ttgggtttca aagctagcac ct 2284 23 DNA Artificial Sequence primer that hybridizes to the human MATN3gene 84 aagtcatcca tacgctaaat cca 23 85 22 DNA Artificial Sequenceprimer that hybridizes to the human MATN3 gene 85 tccaggagca ataagatggaga 22 86 20 DNA Artificial Sequence primer that hybridizes to the humanMATN3 gene 86 ttgctctctg ggctcctttc 20 87 22 DNA Artificial Sequenceprimer that hybridizes to the human MATN3 gene 87 ggaacctatg ctccaggttttg 22 88 22 DNA Artificial Sequence primer that hybridizes to the humanMATN3 gene 88 ttgtagagat ggggtctcac ca 22 89 20 DNA Artificial Sequenceprimer that hybridizes to the human MATN3 gene 89 gttacttggg aggccgagat20 90 20 DNA Artificial Sequence primer that hybridizes to the humanMATN3 gene 90 cccattcctc attgactcca 20 91 22 DNA Artificial Sequenceprimer that hybridizes to the human MATN3 gene 91 agaggggtac tcactggtctgg 22 92 19 DNA Artificial Sequence primer that hybridizes to the humanMATN3 gene 92 cagctgggag ttgggaacc 19 93 20 DNA Artificial Sequenceprimer that hybridizes to the human MATN3 gene 93 tacctacccg gtgatggaag20 94 23 DNA Artificial Sequence primer that hybridizes to the humanMATN3 gene 94 tcacgtttct gagaagtact gga 23 95 24 DNA Artificial Sequenceprimer that hybridizes to the human MATN3 gene 95 tgaggaaaga ttagaaaaacctga 24 96 22 DNA Artificial Sequence primer that hybridizes to thehuman MATN3 gene 96 tcattcaatg cactttcctc tc 22 97 22 DNA ArtificialSequence primer that hybridizes to the human MATN3 gene 97 cactgtttttgcaacctttt ca 22 98 22 DNA Artificial Sequence primer that hybridizes tothe human MATN3 gene 98 tctgaccttc cacaattgct tt 22 99 20 DNA ArtificialSequence primer that hybridizes to the human MATN3 gene 99 aacttgcaactcaccagcaa 20 100 20 DNA Artificial Sequence primer that hybridizes tothe human MATN3 gene 100 catgccattt ccctttgtct 20 101 23 DNA ArtificialSequence primer that hybridizes to the human MATN3 gene 101 aaaaataccggacacctact ttg 23 102 20 DNA Artificial Sequence primer that hybridizesto the human MATN3 gene 102 tgggagtgtt gtctgcctta 20 103 20 DNAArtificial Sequence primer that hybridizes to the human MATN3 gene 103gacctgggaa aacaaaccaa 20 104 21 DNA Artificial Sequence primer thathybridizes to the human MATN3 gene 104 caaaatgtca aaaccccatc t 21 105 24DNA Artificial Sequence primer that hybridizes to the human MATN3 gene105 tgtaccttaa tactcacagc aacg 24 106 22 DNA Artificial Sequence primerthat hybridizes to the human MATN3 gene 106 catggtatgt tccccagctt ta 22107 20 DNA Artificial Sequence primer that hybridizes to the human MATN3gene 107 aagcaataca cctgccttgg 20 108 20 DNA Artificial Sequence primerthat hybridizes to the human MATN3 gene 108 taggttcctc ctctgccatt 20 10922 DNA Artificial Sequence primer that hybridizes to the human MATN3gene 109 agcgtctcac atttcacaag aa 22 110 20 DNA Artificial Sequenceprimer that hybridizes to the human MATN3 gene 110 gatgctgcac aaaaagcact20 111 21 DNA Artificial Sequence primer that hybridizes to the humanMATN3 gene 111 cccatctgtg cttgtgtgtg t 21 112 21 DNA Artificial Sequenceprimer that hybridizes to the human MATN3 gene 112 gaggagaagc ttccatttggt 21 113 22 DNA Artificial Sequence primer that hybridizes to the humanMATN3 gene 113 tcagcccttt aaggctattt cc 22 114 21 DNA ArtificialSequence primer that hybridizes to the human MATN3 gene 114 gagggtgggaatgagtgagt g 21 115 22 DNA Artificial Sequence primer that hybridizes tothe human MATN3 gene 115 cccagattca ctgcttcttc ct 22 116 22 DNAArtificial Sequence primer that hybridizes to the human MATN3 gene 116gcatgggttc agtgggatag tc 22 117 22 DNA Artificial Sequence primer thathybridizes to the human MATN3 gene 117 cccaggtttg agaaatggag tt 22 11822 DNA Artificial Sequence primer that hybridizes to the human MATN3gene 118 tggaattcag tcaggcagct at 22 119 22 DNA Artificial Sequenceprimer that hybridizes to the human MATN3 gene 119 ccagtcttca gacctgggaagt 22 120 22 DNA Artificial Sequence primer that hybridizes to the humanMATN3 gene 120 tgaagtcagg agttcaagac ca 22 121 22 DNA ArtificialSequence primer that hybridizes to the human MATN3 gene 121 acggggtttcactatgttgt cc 22 122 22 DNA Artificial Sequence primer that hybridizesto the human MATN3 gene 122 gaggcagaag gatcacttga gg 22 123 18 DNAArtificial Sequence primer that hybridizes to the human MATN3 gene 123acaggcatgg gccaccac 18 124 22 DNA Artificial Sequence primer thathybridizes to the human MATN3 gene 124 ctggtgcagt gccttattcc ta 22 12522 DNA Artificial Sequence primer that hybridizes to the human MATN3gene 125 tgccactatg cctggctaac ta 22 126 20 DNA Artificial Sequenceprimer that hybridizes to the human MATN3 gene 126 tcctcagtgg ctgaagacaa20 127 22 DNA Artificial Sequence primer that hybridizes to the humanMATN3 gene 127 aagacattcc acaggcaagt ga 22 128 22 DNA ArtificialSequence primer that hybridizes to the human MATN3 gene 128 ccactcctctggggtacata gg 22 129 22 DNA Artificial Sequence primer that hybridizesto the human MATN3 gene 129 gccaagtggt caaaaagcag at 22 130 22 DNAArtificial Sequence primer that hybridizes to the human MATN3 gene 130tgttttctca ccaaaagttg aa 22 131 25 DNA Artificial Sequence primer thathybridizes to the human MATN3 gene 131 atcaaaagac agaagtttga ttctc 25132 22 DNA Artificial Sequence primer that hybridizes to the human MATN3gene 132 cccaggagtt tccaataaac tg 22

What is claimed is:
 1. An isolated nucleic acid molecule comprising amatrilin-3 gene, or a fragment or variant thereof, the nucleotidesequence of SEQ ID NO: 1 and comprising at least one polymorphism asshown in Table 3 or FIGS. 5A-5C.
 2. A nucleic acid encoding apolypeptide having an amino acid sequence selected from the groupconsisting of SEQ ID NOs: 2, 3, 4, 5, 6, 7, 8 and
 9. 3. A method forassaying the presence of a first nucleic acid molecule in a sample,comprising contacting said sample with a second nucleic acid moleculecomprising a nucleotide sequence selected from the group consisting ofSEQ ID NO: 1 comprising at least one polymorphism as shown in Table 3 orFIGS. 5A-5C and complements thereof, under high stringency conditions.4. A vector comprising an isolated nucleic acid molecule selected fromthe group consisting of: SEQ ID NO: 1 and comprising at least onepolymorphism shown in Table 3 or FIGS. 5A-5C, and complements thereof,operatively linked to a regulatory sequence.
 5. A recombinant host cellcomprising the vector of claim
 4. 6. A method for producing apolypeptide encoded by an isolated nucleic acid molecule, comprisingculturing the recombinant host cell of claim 5 under conditions suitablefor expression of said nucleic acid molecule.
 7. An isolated polypeptideencoded by a matrilin-3 gene, wherein the matrilin-3 gene has thesequence of SEQ ID NO: 1 comprising at least one polymorphism as shownin Table 3 or FIGS. 5A-5C, or complements thereof.
 8. The isolatedpolypeptide of claim 7, wherein the polypeptide has an amino acidsequence selected from the group consisting of SEQ ID NOs: 2-9.
 9. Afusion protein comprising an isolated polypeptide of claim
 7. 10. Anantibody, or an antigen-binding fragment thereof, which selectivelybinds to a polypeptide of claim
 7. 11. An antibody, or anantigen-binding fragment thereof, which selectively binds to an aminoacid sequence selected from the group consisting of SEQ ID NOs: 2-9, orto a fragment or variant of said amino acid sequence.
 12. A method forassaying the presence of a polypeptide encoded by an isolated nucleicacid molecule according to claim 1 in a sample, comprising contactingsaid sample with an antibody which specifically binds to the encodedpolypeptide.
 13. A method of diagnosing a susceptibility toosteoarthritis in an individual, comprising detecting a polymorphism ina matrilin-3 gene, wherein the presence of the polymorphism in the geneis indicative of a susceptibility to osteoarthritis.
 14. A method ofdiagnosing a susceptibility to osteoarthritis, comprising detecting analteration in the expression or composition of a polypeptide encoded bya matrilin-3 gene in a test sample, in comparison with the expression orcomposition of a polypeptide encoded by matrilin-3 gene in a controlsample, wherein the presence of an alteration in expression orcomposition of the polypeptide in the test sample is indicative of asusceptibility to osteoarthritis.
 15. The method of claim 14, whereinthe alteration in the expression or composition of a polypeptide encodedby matrilin-3 gene comprises expression of a splicing variantpolypeptide in a test sample that differs from a splicing variantpolypeptide expressed in a control sample.
 16. A method of identifyingan agent which alters activity of a polypeptide of claim 7, comprising:a) contacting the polypeptide or a derivative or fragment thereof, withan agent to be tested; b) assessing the level of activity of thepolypeptide or derivative or fragment thereof; and c) comparing thelevel of activity with a level of activity of the polypeptide or activederivative or fragment thereof in the absence of the agent, wherein ifthe level of activity of the polypeptide or derivative or fragmentthereof in the presence of the agent differs, by an amount that isstatistically significant, from the level in the absence of the agent,then the agent is an agent that alters activity of the polypeptide. 17.A method of identifying an agent which alters interaction of thepolypeptide of claim 7 with a matrilin-3 gene binding agent, comprising:a) contacting the polypeptide or a derivative or fragment thereof, thebinding agent and with an agent to be tested; b) assessing theinteraction of the polypeptide or derivative or fragment thereof withthe binding agent; and c) comparing the level of interaction with alevel of interaction of the polypeptide or derivative or fragmentthereof with the binding agent in the absence of the agent, wherein ifthe level of interaction of the polypeptide or derivative or fragmentthereof in the presence of the agent differs, by an amount that isstatistically significant, from the level of interaction in the absenceof the agent, then the agent is an agent that alters interaction of thepolypeptide with the binding agent.
 18. An agent which altersinteraction of a matrilin-3 gene polypeptide with a matrilin-3 genebinding agent, identifiable according to the method of claim
 17. 19. Amethod of identifying an agent which alters expression of a matrilin-3gene, comprising the steps of: a) contacting a solution containing anucleic acid of claim 1 or a derivative or fragment thereof with anagent to be tested; b) assessing the level of expression of the nucleicacid, derivative or fragment; and c) comparing the level of expressionwith a level of expression of the nucleic acid, derivative or fragmentin the absence of the agent, wherein if the level of expression of thenucleotide, derivative or fragment in the presence of the agent differs,by an amount that is statistically significant, from the expression inthe absence of the agent, then the agent is an agent that altersexpression of a matrilin-3 gene.
 20. A method of identifying an agentwhich alters expression of a matrilin-3 gene, comprising the steps of:a) contacting a solution containing a nucleic acid comprising thepromoter region of matrilin-3 gene operably linked to a reporter gene,with an agent to be tested; b) assessing the level of expression of thereporter gene; and c) comparing the level of expression with a level ofexpression of the reporter gene in the absence of the agent, wherein ifthe level of expression of the reporter gene in the presence of theagent differs, by an amount that is statistically significant, from thelevel of expression in the absence of the agent, then the agent is anagent that alters expression of a matrilin-3 gene.
 21. A method ofidentifying an agent which alters expression of a matrilin-3 gene,comprising the steps of: a) contacting a solution containing a nucleicacid of claim 1 or a derivative or fragment thereof with an agent to betested; b) assessing expression of the nucleic acid, derivative orfragment; and c) comparing expression with expression of the nucleicacid, derivative or fragment in the absence of the agent, wherein ifexpression of the nucleotide, derivative or fragment in the presence ofthe agent differs, by an amount that is statistically significant, fromthe expression in the absence of the agent, then the agent is an agentthat alters expression of a matrilin-3 gene.
 22. The method of claim 21,wherein the expression of the nucleotide, derivative or fragment in thepresence of the agent comprises expression of one or more splicingvariant(s) that differ in kind or in quantity from the expression of oneor more splicing variant(s) the absence of the agent.
 23. A method ofaltering expression of a matrilin-3 gene, comprising contacting a cellcontaining said matrilin-3 gene with an agent identifiable by the methodof claim
 21. 24. A method of identifying a polypeptide which interactswith a matrilin-3 gene polypeptide, comprising employing a yeasttwo-hybrid system using a first vector which comprises a nucleic acidencoding a DNA binding domain and a matrilin-3 gene polypeptide,splicing variant, fragment or derivative thereof, and a second vectorwhich comprises a nucleic acid encoding a transcription activationdomain and a nucleic acid encoding a test polypeptide, wherein iftranscriptional activation occurs in the yeast two-hybrid system, thetest polypeptide is a polypeptide which interacts with a matrilin-3polypeptide.
 25. A matrilin-3 gene therapeutic agent selected from thegroup consisting of: a matrilin-3 gene or fragment or derivativethereof; a polypeptide encoded by matrilin-3 gene; a matrilin-3 genereceptor; a matrilin-3 gene binding agent; a peptidomimetic; a fusionprotein; a prodrug; an antibody; an agent that alters matrilin-3 geneexpression; an agent that alters activity of a polypeptide encoded bymatrilin-3 gene; an agent that alters posttranscriptional processing ofa polypeptide encoded by matrilin-3 gene; an agent that altersinteraction of a matrilin-3 gene with a matrilin-3 gene binding agent;an agent that alters transcription of splicing variants encoded bymatrilin-3 gene; and a ribozyme.
 26. A pharmaceutical compositioncomprising a matrilin-3 gene therapeutic agent of claim
 25. 27. Thepharmaceutical composition of claim 26, wherein the matrilin-3 genetherapeutic agent is an isolated nucleic acid molecule comprising amatrilin-3 gene or fragment or derivative thereof.
 28. Thepharmaceutical composition of claim 26, wherein the matrilin-3 genetherapeutic agent is a polypeptide encoded by the matrilin-3 gene.
 29. Amethod of treating osteoarthritis in an individual, comprisingadministering a matrilin-3 gene therapeutic agent to the individual, ina therapeutically effective amount.
 30. The method of claim 29, whereinthe matrilin-3 gene therapeutic agent is a matrilin-3 gene agonist. 31.The method of claim 30, wherein the matrilin-3 gene therapeutic agent isa matrilin-3 gene antagonist.
 32. A transgenic animal comprising anucleic acid selected from the group consisting of: an exogenousmatrilin-3 gene and a nucleic acid encoding a matrilin-3 genepolypeptide.
 33. A method for assaying a sample for the presence of amatrilin-3 gene nucleic acid, comprising: a) contacting said sample witha nucleic acid comprising a contiguous nucleotide sequence, which is aat least partially complementary to a part of the sequence of saidmatrilin-3 gene nucleic acid under conditions suitable forhybridization, and b) assessing whether hybridization has occurredbetween a matrilin-3 gene nucleic acid and said nucleic acid comprisinga contiguous nucleotide sequence which is at least partiallycomplementary to a part of the sequence of said matrilin-3 gene nucleicacid.
 34. The method of claim 33, wherein said nucleic acid comprising acontiguous nucleotide sequence is completely complementary to a part ofthe sequence of said matrilin-3 gene nucleic acid.
 35. The method ofclaim 33, comprising amplification of at least part of said matrilin-3gene nucleic acid.
 36. The method of claim 33, wherein said contiguousnucleotide sequence is 100 or fewer nucleotides in length and is either:a) at least 80% identical to a contiguous sequence of nucleotides in SEQID NO: 1 comprising at least one polymorphism as shown in Table 3 orFIGS. 5A-5C; b) at least 80% identical to the complement of a contiguoussequence of nucleotides in SEQ ID NO: 1 comprising at least onepolymorphism as shown in Table 3 or FIGS. 5A-5C; or c) capable ofselectively hybridizing to said matrilin-3 gene nucleic acid.
 37. Areagent for assaying a sample for the presence of a matrilin-3 genenucleic acid, said reagent comprising a nucleic acid comprising acontiguous nucleotide sequence which is at least partially complementaryto a part of the nucleotide sequence of said matrilin-3 gene nucleicacid.
 38. The reagent of claim 37, wherein the nucleic acid comprises acontiguous nucleotide sequence which is completely complementary to apart of the nucleotide sequence of said matrilin-3 gene nucleic acid.39. A reagent kit for assaying a sample for the presence of a matrilin-3gene nucleic acid comprising in separate containers: a) one or morelabeled nucleic acids comprising a contiguous nucleotide sequence whichis at least partially complementary to a part of the nucleotide sequenceof said matrilin-3 gene nucleic acid; and b) reagents for detection ofsaid label.
 40. The reagent kit of claim 39, wherein the labeled nucleicacid comprises a contiguous nucleotide sequences which is completelycomplementary to a part of the nucleotide sequence of said matrilin-3gene nucleic acid.
 41. A reagent kit for assaying a sample for thepresence of a matrilin-3 gene nucleic acid comprising one or morenucleic acids comprising a contiguous nucleotide sequence which is atleast partially complementary to a part of the nucleotide sequence ofsaid matrilin-3 gene nucleic acid, and which is capable of acting as aprimer for said matrilin-3 gene nucleic acid when maintained underconditions for primer extension.
 42. A method of diagnosingsusceptibility to osteoarthritis in an individual, comprising screeningfor an at-risk haplotype in the matrilin-3 gene that is more frequentlypresent in an individual susceptible to osteoarthritis compared to ahealthy individual, wherein the at-risk haplotype increases risk ofosteoarthritis significantly.
 43. The method of claim 42, wherein thesignificant increase is at least about 20%.
 44. The method of claim 42,wherein the significant increase is identified as an odds ratio of atleast about 1.2.
 45. A method of diagnosing susceptibility toosteoarthritis in an individual, comprising screening for an at-riskhaplotype in the matrilin-3 gene that is more frequently present in anindividual susceptible to osteoarthritis (affected), compared to thefrequency of its presence in a healthy individual (control), wherein thepresence of the at-risk haplotype is indicative of a susceptibility toosteoarthritis.
 46. The method of claim 45, wherein the at-riskhaplotype is characterized by the presence of at least one polymorphismat nucleic acid positions 58162, 57927, 56822, 47929, 45434, 45317,45178 and 45010, relative to SEQ ID NO:
 1. 47. The method of claim 45,wherein screening for the presence of an at-risk haplotype in thematrilin-3 gene comprises enzymatic amplification of nucleic acid fromsaid individual.
 48. The method of claim 45, wherein the nucleic acid isDNA.
 49. The method of claim 48, wherein the DNA is mammalian.
 50. Themethod of claim 49, wherein the DNA is human.
 51. The method of claim45, wherein screening for the presence of an at-risk haplotype in thematrilin-3 gene comprises: (a) obtaining material containing nucleicacid from the individual; (b) amplifying said nucleic acid; and (c)determining the presence or absence of an at-risk haplotype in saidamplified nucleic acid.
 52. The method of claim 51, wherein determiningthe presence of an at-risk haplotype is performed by electrophoreticanalysis.
 53. The method of claim 51, wherein determining the presenceof an at-risk haplotype is performed by restriction length polymorphismanalysis.
 54. The method of claim 51, wherein determining the presenceof an at-risk haplotype is performed by sequence analysis.
 55. Themethod of claim 51, wherein determining the presence of an at-riskhaplotype is performed by hybridization analysis.
 56. A kit fordiagnosing susceptibility to osteoarthritis in an individual comprising:primers for nucleic acid amplification of a region of the matrilin-3gene comprising an at-risk haplotype, wherein the primers comprise asegment of nucleic acids of length suitable for nucleic acidamplification, selected from the group consisting of: a polymorphism atnucleic acid position 58162, 57927, 56822, 47929, 45434, 45317, 45178and 45010, relative to SEQ ID NO: 1 and combinations thereof.